Solid Forms and Formulations of Imidazopyrazine Compound

ABSTRACT

In some embodiments, the invention relates to crystalline solid forms, including hydrates, polymorphs, and salt forms, of (S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide. In some embodiments, the invention relates to amorphous solid forms of (S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide. In some embodiments, the invention also relates to pharmaceutical compositions containing the solid forms, and methods for treating conditions or disorders by administering to a subject a pharmaceutical composition that includes the forms, including pharmaceutical compositions and methods for overcoming the effects of acid reducing agents.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims priority to, and the benefit of, U.S.Provisional Application No. 62/188,468, filed on Jul. 2, 2015, and U.S.Provisional Application No. 62/271,708, filed on Dec. 28, 2015, theentirety of each of which is incorporated herein by reference.

FIELD OF THE INVENTION

In some embodiments, the invention relates to crystalline Form I of(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide.In other embodiments, the invention relates to pharmaceuticalcompositions including Form I, including pharmaceutical compositionsthat overcome the effects of acid reducing agents, and methods fortreating cancers or other disorders by administering the pharmaceuticalcompositions to a subject. In some embodiments, the invention relates tocrystalline salts of(5)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide.In other embodiments, the invention relates to pharmaceuticalcompositions including crystalline salts of(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide,including pharmaceutical compositions that overcome the effects of acidreducing agents, and methods for treating cancers or other disorders byadministering the pharmaceutical compositions to a subject.

BACKGROUND OF THE INVENTION

Bruton's Tyrosine Kinase (BTK) is a Tec family non-receptor proteinkinase expressed in B cells and myeloid cells. BTK is composed of thepleckstrin homology (PH), Tec homology (TH), Src homology 3 (SH3), Srchomology 2 (SH2), and tyrosine kinase or Src homology 1 (TK or SH1)domains. The function of BTK in signaling pathways activated by theengagement of the B cell receptor (BCR) in mature B cells and FCER1 onmast cells is well established. Functional mutations in BTK in humansresult in a primary immunodeficiency disease (X-linkedagammaglobuinaemia) characterized by a defect in B cell development witha block between pro- and pre-B cell stages. The result is an almostcomplete absence of B lymphocytes, causing a pronounced reduction ofserum immunoglobulin of all classes. These findings support a key rolefor BTK in the regulation of the production of auto-antibodies inautoimmune diseases. BTK is expressed in numerous B cell lymphomas andleukemias. Other diseases with an important role for dysfunctional Bcells are B cell malignancies, as described in Hendriks, et al., Nat.Rev. Cancer, 2014, 14, 219-231. The reported role for BTK in theregulation of proliferation and apoptosis of B cells indicates thepotential for BTK inhibitors in the treatment of B cell lymphomas. BTKinhibitors have thus been developed as potential therapies for many ofthese malignancies, as described in D'Cruz, et al., OncoTargets andTherapy 2013, 6, 161-176; and International Patent ApplicationPublication No. WO 2013/010868 discloses BTK inhibitors including(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefor use in therapy.

The present invention includes the unexpected discovery of novel solidforms of(5)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide,referred to herein as Formula (1). Formula (1) is a BTK inhibitor thatis useful, inter alia, in pharmaceutical compositions and methods fortreatment of cancers, inflammation, immune, and autoimmune diseases. Thenovel solid forms of Formula (1) disclosed herein have surprising anduseful properties.

SUMMARY OF THE INVENTION

In an embodiment, the invention provides a composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base.

In an embodiment, the invention provides a composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by an X-ray powder diffraction patterncomprising peaks at 6.4, 8.6, 10.5, 11.6, and 15.7 °2θ±0.2 °2θ.

In an embodiment, the invention provides a composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by an X-ray powder diffraction patterncomprising peaks at 6.4, 8.6, 10.5, 11.6, and 15.7 °2θ±0.2 °2θ andfurther comprising peaks at 10.9, 12.7, 13.4, 14.3, 14.9, and 18.2°2θ±0.2 °2θ.

In an embodiment, the invention provides a composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by an X-ray powder diffraction patterncomprising peaks at 6.4, 8.6, 10.5, 11.6, and 15.7 °2θ±0.2 °2θ, furthercomprising peaks at 10.9, 12.7, 13.4, 14.3, 14.9, and 18.2 °2θ±0.2 °2θand further comprising peaks selected from the group consisting of 11.3,15.1, 15.7, 16.1, 17.3, 19.2, 19.4, 19.8, 20.7, 21.1, 21.4, 21.6, 21.9,22.6, 23.3, 23.6, 24.9, 25.2, 25.4, 25.7, 26.1, 26.4, 26.8, 26.9, 27.7,28.6, 29.1, 29.4, 30.1, 30.5, 31.7, 31.9, 32.2, 32.6, 33.1, 33.4, 34.5,35.9, 36.1, 36.8, 37.4, 38.1, 38.9, and 39.5 °2θ±0.2 °2θ, and anycombination of one or more peaks thereof.

In an embodiment, the invention provides a composition comprisingcrystalline(5)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by a transmission X-ray powder diffractionpattern substantially the same as the representative X-ray powderdiffraction pattern shown in FIG. 2.

In an embodiment, the invention provides a composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is further characterized by a Raman spectrum comprising peaksat 1620, 1609, 1547, 1514 and 1495 cm⁻¹±4 cm⁻¹.

In an embodiment, the invention provides a composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by a Raman spectrum comprising peaks at 1620,1609, 1547, 1514 and 1495 cm⁻¹±4 cm⁻¹ and further comprising one or morepeaks selected from the group consisting of 1680, 1620, 1609, 1574,1547, 1514, 1495, 1454, 1433, 1351, 1312, 1255, 1232, 1187, 1046, 995,706, 406, 280, and any combination of one or more peaks thereof, withpeak positions measured in cm⁻¹±4 cm⁻¹.

In an embodiment, the invention provides a composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is further characterized by an infrared (IR) spectrumcomprising peaks at 1621, 1608, 1403, 1303, and 764 cm⁻¹±4 cm⁻¹.

In an embodiment, the invention provides a composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by an IR spectrum comprising 1621, 1608,1403, 1303, and 764 cm⁻¹±4 cm⁻¹ and comprising one or more peaksselected from the group consisting of 3367, 3089, 2246, 1682, 1621,1608, 1574, 1514, 1504, 1454, 1428, 1403, 1345, 1303, 1248, 1194, 1177,1149, 1109, 1049, 1023, 1003, 947, 900, 858, 842, 816, 764, 734, 729,701, 689, 665, 623, 612, and any combination of one or more peaksthereof, with peak positions measured in cm⁻¹±4 cm⁻¹.

In an embodiment, the invention provides a composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is further characterized by the absence of water in thecrystal structure.

In an embodiment, the invention provides a composition comprising anextragranular acidulant.

In an embodiment, the invention provides a composition whereinextragranular acidulant is selected from the group consisting of fumaricacid, succinic acid, D-tartaric acid, L-tartaric acid, racemic tartaricacid, ascorbic acid, isoascorbic acid (also known as erythorbic acid andD-araboascorbic acid), alginic acid, Protacid F 120 NM, Protacid AR 1112(also known as Kelacid NF), and Carbopol 971P (carboxypolymethylene),and combinations thereof.

In an embodiment, the invention provides a composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base and an extragranular acidulant, wherein the extragranularacidulant is alginic acid, or a sodium or potassium salt thereof, at aconcentration of between about 5% to about 33% by weight.

In an embodiment, the invention provides a composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base and an extragranular acidulant, wherein the extragranularacidulant is alginic acid, or a sodium or potassium salt thereof, at aconcentration of between about 5% to about 33% by weight, and whereinthe composition further comprises at least onepharmaceutically-acceptable excipient.

In an embodiment, the invention provides a method of treating ahyperproliferative disease comprising the step of administering atherapeutically effective amount of a composition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base Form I to a mammal, wherein the hyperproliferative disease isselected from the group consisting of chronic lymphocytic leukemia,non-Hodgkin's lymphoma, diffuse large B-cell lymphoma, mantle celllymphoma, follicular lymphoma, B-cell lymphoproliferative disease, Bcell acute lymphoblastic leukemia, Waldenström's macroglobulinemia,Burkitt's leukemia, Hodgkin's disease, multiple myeloma, acute myeloidleukemia, juvenile myelomonocytic leukemia, hairy cell leukemia, mastcell leukemia, mastocytosis, myeloproliferative disorders (MPDs),myeloproliferative neoplasms, polycythemia vera (PV), essentialthrombocythemia (ET), primary myelofibrosis (PMF), myelodysplasticsyndrome, chronic myelogenous leukemia (BCR-ABL1-positive), chronicneutrophilic leukemia, chronic eosinophilic leukemia, primary centralnervous system (CNS) lymphoma, primary multifocal lymphoma of peripheralnervous system (PNS), thymus cancer, brain cancer, glioblastoma, lungcancer, squamous cell cancer, skin cancer (e.g., melanoma), eye cancer,retinoblastoma, intraocular melanoma, oral cavity and oropharyngealcancers, bladder cancer, gastric cancer, stomach cancer, pancreaticcancer, breast cancer, cervical cancer, head and neck cancer, renalcancer, kidney cancer, liver cancer, ovarian cancer, prostate cancer,colorectal cancer, bone cancer (e.g., metastatic bone cancer),esophageal cancer, testicular cancer, gynecological cancer, thyroidcancer, epidermoid cancer, AIDS-related cancer (e.g., lymphoma),viral-induced cervical carcinoma (human papillomavirus), nasopharyngealcarcinoma (Epstein-Barr virus), Kaposi's sarcoma, primary effusionlymphoma (Kaposi's sarcoma herpesvirus), hepatocellular carcinoma(hepatitis B and hepatitis C viruses), T-cell leukemias (Human T-cellleukemia virus-1), benign hyperplasia of the skin, restenosis, benignprostatic hypertrophy, tumor angiogenesis, chronic inflammatory disease,rheumatoid arthritis, atherosclerosis, inflammatory bowel disease, skindiseases such as psoriasis, eczema, and scleroderma, diabetes, diabeticretinopathy, retinopathy of prematurity, age-related maculardegeneration, hemangioma, ulcerative colitis, atopic dermatitis,pouchitis, spondylarthritis, uveitis, Behcet's disease, polymyalgiarheumatica, giant-cell arteritis, sarcoidosis, Kawasaki disease,juvenile idiopathic arthritis, hidratenitis suppurativa, Sjögren'ssyndrome, psoriatic arthritis, juvenile rheumatoid arthritis, ankylosingspondylitis, Crohn's disease, lupus, and lupus nephritis.

In an embodiment, the invention provides a method of treating ahyperproliferative disease comprising the step of administering atherapeutically effective amount of a composition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base Form I to a mammal, wherein the hyperproliferative disease isselected from the group consisting of chronic lymphocytic leukemia,non-Hodgkin's lymphoma, diffuse large B-cell lymphoma, mantle celllymphoma, follicular lymphoma, and Waldenström's macroglobulinemia.

In an embodiment, the invention provides a method of treating ahyperproliferative disease comprising the step of administering atherapeutically effective amount of a composition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base Form I and an extragranular acidulant to a mammal, wherein thehyperproliferative disease is selected from the group consisting ofchronic lymphocytic leukemia, non-Hodgkin's lymphoma, diffuse largeB-cell lymphoma, mantle cell lymphoma, follicular lymphoma, andWaldenström's macroglobulinemia.

In an embodiment, the invention provides a method of treating ahyperproliferative disease comprising the step of administering atherapeutically effective amount of a composition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base Form I and an extragranular acidulant to a mammal, furthercomprising the step of administering a therapeutically effective amountof an acid-reducing agent to the mammal.

In an embodiment, the invention provides a method of treating ahyperproliferative disease comprising the step of administering atherapeutically effective amount of a composition comprising acrystalline fumarate, maleate, phosphate, L-tartrate, citrate,gentisate, oxalate, or sulfate salt of(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base Form I to a mammal, further comprising the step ofadministering a therapeutically effective amount of an acid-reducingagent to the mammal.

BRIEF DESCRIPTION OF THE DRAWINGS

The foregoing summary, as well as the following detailed description ofthe invention, will be better understood when read in conjunction withthe appended drawings.

FIG. 1 illustrates a powder X-ray diffraction (PXRD) pattern of Form I(sample PP502-P1) of the free base of Formula (1) measured in reflectionmode.

FIG. 2 illustrates a PXRD pattern of Form I (sample PP502-P1) of thefree base of Formula (1) measured in transmission mode.

FIG. 3 illustrates optical polarized microscope images of Form I (samplePP502-P1) of the free base of Formula (1). The left image showsundispersed crystals. The right image shows crystals dispersed withparaffin oil.

FIG. 4 illustrates a Raman spectrum of Form I of the free base ofFormula (1) in the spectral range from 200 to 1800 cm⁻¹.

FIG. 5 illustrates an infrared (IR) spectrum of Form I of the free baseof Formula (1).

FIG. 6 illustrates an IR spectrum of Form I of the free base of Formula(1).

FIG. 7 illustrates a ¹H nuclear magnetic resonance (NMR) spectrum ofForm I of the free base of Formula after dissolution in d₆-DMSO.

FIG. 8 illustrates a thermogram from thermogravimetric analysis (TGA)with Fourier transform IR spectroscopy (TG-FTIR) of Form I of the freebase of Formula (1) (PP502-P1).

FIG. 9 illustrates a thermogram from differential scanning calorimetry(DSC) analysis of Form I of the free base of Formula (1) (PP502-P1).

FIG. 10 illustrates a plot of the water content vs. time and relativehumidity (RH) for Form I of the free base of Formula (1). The red lineis the water content (left y-axis) and the blue line reflects themeasurement program to which the sample was exposed (right y-axis).

FIG. 11 illustrates a dynamic vapor sorption (DVS) isotherm plot withwater content versus RH for Form I of the free base of Formula (1).

FIG. 12 illustrates a PXRD pattern of Form II of the free base ofFormula (1).

FIG. 13 illustrates a polarized microscope image of Form II of the freebase of Formula (1) dispersed in paraffin oil.

FIG. 14 illustrates a Raman spectrum of Form II of the free base ofFormula (1) in the spectral range from 200 to 1800 cm⁻¹.

FIG. 15 illustrates an IR spectrum of Form II of the free base ofFormula (1).

FIG. 16 illustrates an IR spectrum of Form II of the free base ofFormula (1).

FIG. 17 illustrates a TGA thermogram from TG-FTIR analysis of Form II ofthe free base of Formula (1).

FIG. 18 illustrates a DSC thermogram of Form II of the free base ofFormula (1).

FIG. 19 illustrates a plot of the water content versus time and RH forForm II of the free base of Formula (1).

FIG. 20 illustrates an isotherm plot of the DVS analysis for Form II ofthe free base of Formula (1).

FIG. 21 illustrates a PXRD pattern of Form III of the free base ofFormula (1).

FIG. 22 illustrates a Raman spectrum of Form III.

FIG. 23 illustrates an IR spectrum of Form III of the free base ofFormula (1).

FIG. 24 illustrates an IR spectrum of Form III of the free base ofFormula (1).

FIG. 25 illustrates a DSC thermogram of Form III of the free base ofFormula (1).

FIG. 26 illustrates a PXRD pattern of Form IV of the free base ofFormula (1) (Form II after dehydration under nitrogen).

FIG. 27 illustrates a PXRD pattern of Form V of the free base of Formula(1).

FIG. 28 illustrates a Raman spectrum of Form V of the free base ofFormula (1) in the spectral range from 200 to 1800 cm⁻¹.

FIG. 29 illustrates a PXRD pattern of Form VI.

FIG. 30 illustrates a Raman spectrum of Form VI in the spectral rangefrom 200 to 1800 cm⁻¹.

FIG. 31 illustrates a PXRD pattern of Form VII.

FIG. 32 illustrates a Raman spectrum of Form VII in the spectral rangefrom 200 to 1800 cm⁻¹.

FIG. 33 illustrates a PXRD pattern of Form VIII.

FIG. 34 illustrates a Raman spectrum of Form VIII in the spectral rangefrom 200 to 1800 cm⁻¹.

FIG. 35 illustrates the interrelationship between several forms ofFormula (1).

FIG. 36 illustrates the PXRD pattern of a sample of amorphous Formula(1).

FIG. 37 illustrates the Raman spectrum of a sample of amorphous Formula(1).

FIG. 38 illustrates the IR spectrum of a sample of amorphous Formula(1).

FIG. 39 illustrates a FTIR thermogram of a sample of amorphous Formula(1).

FIG. 40 illustrates a DSC thermogram of a sample of amorphous (Formula)(1).

FIG. 41 illustrates PXRD patterns of the fumarate salt of Formula (1).

FIG. 42 illustrates a PXRD pattern of Formula (1) maleate salt ofFormula (1).

FIG. 43 illustrates a PXRD pattern of the phosphate salt of Formula (1).

FIG. 44 illustrates a PXRD pattern of the L-tartrate salt of Formula(1).

FIG. 45 illustrates a species distribution for Formula (1) based oncalculated pH values: 2.2, 6.1 and 11.5.

FIG. 46 illustrates the pH-dependent solubility of the free base ofFormula (1) with HCl and buffer solutions as the solvent media.

FIG. 47 illustrates the species distribution and solubility as afunction of pH for Formula (1).

FIG. 48 illustrates the temperature dependent solubility of Formula (1).

FIG. 49 illustrates a PXRD pattern of Form I of the free base of Formula(1) recrystallized from ethanol.

FIG. 50 illustrates a temperature cycling profile for suspensionequilibration experiments PP502-P55 through PP502-P58.

FIG. 51 illustrates light microscope images from samples of experimentsPP502-P55 through PP502-P58 (left two columns) and the starting material(sample PP502-P1, right column). Recorded with crossed polarizers anddispersed in paraffin oil.

FIG. 52 illustrates optical microscopy images from samples ofexperiments PP502-P75 through PP502-P77. The images were recorded withcrossed polarizers dispersed in paraffin oil.

FIG. 53 illustrates the labeling scheme and molecular conformationobtained from the single-crystal X-ray diffraction (SCXRD) study of FormI for molecules A (top) and B (bottom), with non-hydrogen atomsrepresented as thermal ellipsoids drawn at the 50% probability level.Hydrogen atoms are shown as spheres of arbitrary size.

FIG. 54 compares the PXRD pattern of Form I to the pattern simulatedusing the crystal structure of Form I.

FIG. 55 shows intrinsic dissolution rate results for Forms I and II ofFormula (1) free base.

FIG. 56 shows exposure data in dogs for Forms I and II of Formula (1)free base.

FIG. 57 illustrates a PXRD pattern of a sample of the L-arabitolcocrystal of Formula (1).

FIG. 58 illustrates a PXRD pattern of Form A of the citrate salt ofFormula (1) (sample SP211-CIT-P4) crystallized from acetone-water.

FIG. 59 illustrates a Raman spectrum of a sample of Form A of thecitrate salt of Formula (1).

FIG. 60 illustrates a PXRD pattern of a sample of Form A of thegentisate salt of Formula (1) monohydrate.

FIG. 61 illustrates a PXRD pattern of Form A of the oxalate salt ofFormula (1).

FIG. 62 illustrates PXRD patterns of materials likely to be other formsof oxalate salts of Formula (1).

FIG. 63 illustrates PXRD patterns of Formula (1) L-proline cocrystalsamples.

FIG. 64 illustrates a PXRD pattern of Form A of the D-sorbitol cocrystalof Formula (1).

FIG. 65 illustrates a PXRD pattern of Form A of the succinic acidcocrystal of Formula (1).

FIG. 66 illustrates a PXRD pattern of a sample of Form A of a sulfatesalt of Formula (1).

FIG. 67 illustrates a comparison of the dissolution profiles offormulations of Formula (1) at a pH of 3.4.

FIG. 68 illustrates a comparison of the dissolution profiles offormulations of Formula (1) at a pH of 5.5.

FIG. 69 shows individual concentration-time profiles from dogs in theomeprazole/Formula (1) study. Conditioned dogs were dosed with 100 mg ofFormula (1) in sequential dosing phases separated by washout periods of4-7 days. Liquid capsules or solid capsules containing Form I of Formula(1) were administered; for comparison, a clinical formulation orhand-filled capsules with an Avicel blend were administered. After theseinitial study phases, dogs were treated with 10 mg/day omeprazolethroughout the remainder of the study. After four days of omeprazoletreatment, 100 mg of Form I of Formula (1) was administered in theclinical formulation, in a formulation containing acidulants, or incapsules containing 100 mg free base equivalent of the Formula (1) saltsof maleate, phosphate, fumarate or tartrate, and plasma concentrationsof Formula (1) were measured at the indicated times.

FIG. 70 shows trends in the AUC, C_(max), and T_(max) for conditioneddogs treated with various solid forms of Formula (1). Liquid capsules(100 mg) were administered for comparison with the solid forms. Solidcapsules of 100 mg strength in the clinical formulation of Form I ofFormula (1) were administered to dogs prior to or following dailytreatment with omeprazole to reduce stomach acidity. The subsequentstudy phases followed 4 days of dosing with omeprazole (10 mg/day);omeprazole treatment continued throughout the study. An acidulantformulation of Form I of Formula (1) was compared with the maleate,phosphate, fumarate or tartrate salts of Formulation 1 (F-1)administered as 100 mg equivalent of free base in capsules. Exposures ofsalts and the capsules of Form I of Formula (1) formulated withacidulant were increased, relative to exposure of Form I in the presenceof omeprazole.

FIG. 71 illustrates dose-normalized AUC and C_(max) for Formula (1) indogs, comparing liquid capsules (“Liq Caps”) (average of n=2),formulation F-2 (average of n=5), formulation F-2 with omeprazole(“F-2/Omep,” showing loss of exposure for Formula (1)), and fiveformulations of the present invention that restore exposure in thepresence of omeprazole: FA-3 (with acidulant, “FA-3/Omep”), F-1 maleate(“Maleate/Omep”), F-1 phosphate (“Phosphate/Omep”), F-1 fumarate(“Fumarate/Omep”), and F-1 tartrate (“Tartrate/Omep”).

FIG. 72 shows PXRD patterns for amorphous solid dispersions (samplesPP502-P128, PP502-P129, PP502-P130, PP502-P131, PP502-P132, PP502-P136,PP502-P137, PP502-P138, and PP502-P139).

DETAILED DESCRIPTION OF THE INVENTION

While preferred embodiments of the invention are shown and describedherein, such embodiments are provided by way of example only and are notintended to otherwise limit the scope of the invention. Variousalternatives to the described embodiments of the invention may beemployed in practicing the invention.

Definitions

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as is commonly understood by one of skill in theart to which this invention belongs. All patents and publicationsreferred to herein are incorporated by reference in their entireties.

The term “solid form” may refer to a crystalline solid form or phase,including crystalline free base, crystalline salt, or a cocrystal, aswell as an amorphous phase, including an amorphous dispersion.

The terms “co-administration,” “co-administering,” “administered incombination with,” and “administering in combination with” as usedherein, encompass administration of two or more agents to a subject sothat both agents and/or their metabolites are present in the subject atthe same time. Co-administration includes simultaneous administration inseparate compositions, administration at different times in separatecompositions, or administration in a composition in which two or moreagents are present.

The term “effective amount” or “therapeutically effective amount” refersto that amount of a compound or combination of compounds as describedherein that is sufficient to effect the intended application including,but not limited to, disease treatment. A therapeutically effectiveamount may vary depending upon the intended application (in vitro or invivo), or the subject and disease condition being treated (e.g., theweight, age and gender of the subject), the severity of the diseasecondition, the manner of administration, etc. which can readily bedetermined by one of ordinary skill in the art. The term also applies toa dose that will induce a particular response in target cells (e.g., thereduction of platelet adhesion and/or cell migration). The specific dosewill vary depending on the particular compounds chosen, the dosingregimen to be followed, whether the compound is administered incombination with other compounds, timing of administration, the tissueto which it is administered, and the physical delivery system in whichthe compound is carried.

The terms “QD,” “qd,” or “q.d.” mean quaque die, once a day, or oncedaily. The terms “BID,” “bid,” or “b.i.d.” mean bis in die, twice a day,or twice daily. The terms “TID,” “tid,” or “t.i.d.” mean ter in die,three times a day, or three times daily. The terms “QID,” “qid,” or“q.i.d.” mean quater in die, four times a day, or four times daily.

A “therapeutic effect” as that term is used herein, encompasses atherapeutic benefit and/or a prophylactic benefit as described above. Aprophylactic effect includes delaying or eliminating the appearance of adisease or condition, delaying or eliminating the onset of symptoms of adisease or condition, slowing, halting, or reversing the progression ofa disease or condition, or any combination thereof.

The term “pharmaceutically acceptable salt” refers to salts derived froma variety of organic and inorganic counter ions including fumarate,maleate, phosphate, L-tartrate, citrate, gentisate, oxalate, and sulfatecounter ions. Pharmaceutically acceptable acid addition salts can beformed with inorganic acids and organic acids. “Pharmaceuticallyacceptable carrier” or “pharmaceutically acceptable excipient” isintended to include any and all solvents, dispersion media, coatings,antibacterial and antifungal agents, isotonic and absorption delayingagents. Except insofar as any conventional media or agent isincompatible with the active ingredient, its use in the therapeuticcompositions of the invention is contemplated. Supplementary activeingredients can also be incorporated into the described compositions.

The term “in vivo” refers to an event that takes place in a subject'sbody.

The term “in vitro” refers to an event that takes places outside of asubject's body. In vitro assays encompass cell-based assays in whichcells alive or dead are employed and may also encompass a cell-freeassay in which no intact cells are employed.

The term “extragranular” refers to substances that are outside of agranule, e.g., a substance added to granules (multiparticle compactsformed by a granulation process) and physically mixed with granules, butnot contained within the granules.

The term “intragranular” refers to substances that are within a granule(a multiparticle compact formed by a granulation process). Granules maybe formed by processes such as wet granulation (i.e., prepared usingmoisture or steam, thermal, melt, freeze, foam, and other processes) ordry granulation.

The term “acidulant” refers to a substance that increases acidity.

The terms “transmission” or “transmission mode,” when used inconjunction with powder X-ray diffraction, refers to the transmission(also known as Debye-Scherrer) sampling mode. The terms “reflection” or“reflection mode,” when used in conjunction with powder X-raydiffraction, refers to the reflection (also known as Bragg-Brentano)sampling mode.

The term “amorphous solid molecular dispersion” refers to dispersions ofcompounds such as Formula (1) in, e.g., a polymeric excipient, whereinthe polymer and compound are mixed intimately, e.g., at a molecularlevel or at a nanoscale level.

Unless otherwise stated, the chemical structures depicted herein areintended to include compounds which differ only in the presence of oneor more isotopically enriched atoms. For example, compounds where one ormore hydrogen atoms is replaced by deuterium or tritium, or wherein oneor more carbon atoms is replaced by ¹³C- or ¹⁴C-enriched carbons, arewithin the scope of this invention.

When ranges are used herein to describe, for example, physical orchemical properties such as molecular weight or chemical formulae, allcombinations and subcombinations of ranges and specific embodimentstherein are intended to be included. Use of the term “about” or“approximately” when referring to a number or a numerical range meansthat the number or numerical range referred to is an approximationwithin experimental variability (or within statistical experimentalerror), and thus the number or numerical range may vary from, forexample, between 1% and 15% of the stated number or numerical range. Theterm “comprising” (and related terms such as “comprise” or “comprises”or “having” or “including”) includes those embodiments such as, forexample, an embodiment of any composition of matter, method or processthat “consist of” or “consist essentially of” the described features.

“Enantiomeric purity” as used herein refers to the relative amounts,expressed as a percentage, of the presence of a specific enantiomerrelative to the other enantiomer. For example, if a compound, which maypotentially have an (R)- or an (S)-isomeric configuration, is present asa racemic mixture, the enantiomeric purity is about 50% with respect toeither the (R)- or (S)-isomer. If that compound has one isomeric formpredominant over the other, for example, 80% (S)-isomer and 20%(R)-isomer, the enantiomeric purity of the compound with respect to the(S)-isomeric form is 80%. The enantiomeric purity of a compound can bedetermined in a number of ways, including but not limited tochromatography using a chiral support, polarimetric measurement of therotation of polarized light, nuclear magnetic resonance spectroscopyusing chiral shift reagents which include but are not limited tolanthanide containing chiral complexes or Pirkle's reagents, orderivatization of a compounds using a chiral compound such as Mosher'sacid followed by chromatography or nuclear magnetic resonancespectroscopy.

In preferred embodiments, the enantiomerically enriched composition hasa higher potency with respect to therapeutic utility per unit mass thandoes the racemic mixture of that composition. Enantiomers can beisolated from mixtures by methods known to those skilled in the art,including chiral high pressure liquid chromatography (HPLC) and theformation and crystallization of chiral salts; or preferred enantiomerscan be prepared by asymmetric syntheses. See, for example, Jacques, etal., Enantiomers, Racemates and Resolutions, Wiley Interscience, NewYork, 1981; Eliel, Stereochemistry of Carbon Compounds, McGraw-Hill, NY,1962; and Eliel and Wilen, Stereochemistry of Organic Compounds,Wiley-Interscience, New York, 1994.

The terms “enantiomerically enriched” and “non-racemic,” as used herein,refer to compositions in which the percent by weight of one enantiomeris greater than the amount of that one enantiomer in a control mixtureof the racemic composition (e.g., greater than 1:1 by weight). Forexample, an enantiomerically enriched preparation of the (S)-enantiomer,means a preparation of the compound having greater than 50% by weight ofthe (S)-enantiomer relative to the (R)-enantiomer, such as at least 75%by weight, or such as at least 80% by weight. In some embodiments, theenrichment can be significantly greater than 80% by weight, providing a“substantially enantiomerically enriched” or a “substantiallynon-racemic” preparation, which refers to preparations of compositionswhich have at least 85% by weight of one enantiomer relative to otherenantiomer, such as at least 90% by weight, or such as at least 95% byweight. The terms “enantiomerically pure” or “substantiallyenantiomerically pure” refers to a composition that comprises at least98% of a single enantiomer and less than 2% of the opposite enantiomer.

“Moiety” refers to a specific segment or functional group of a molecule.Chemical moieties are often recognized chemical entities embedded in orappended to a molecule.

“Tautomers” are structurally distinct isomers that interconvert bytautomerization. “Tautomerization” is a form of isomerization andincludes prototropic or proton-shift tautomerization, which isconsidered a subset of acid-base chemistry. “Prototropictautomerization” or “proton-shift tautomerization” involves themigration of a proton accompanied by changes in bond order, often theinterchange of a single bond with an adjacent double bond. Wheretautomerization is possible (e.g., in solution), a chemical equilibriumof tautomers can be reached. An example of tautomerization is keto-enoltautomerization. A specific example of keto-enol tautomerization is theinterconversion of pentane-2,4-dione and 4-hydroxypent-3-en-2-onetautomers. Another example of tautomerization is phenol-ketotautomerization. The formation of solid forms in differenttautomerization states is known as “desmotropy” and such forms are knownas “desmotropes.”

Compositions of the invention also include crystalline forms of thosecompounds, including, for example, polymorphs, pseudopolymorphs,solvates, hydrates, unsolvated polymorphs (including anhydrates), andconformational polymorphs, as well as mixtures thereof “Crystallineform”, “form” and “polymorph” are intended to include all crystallineforms of the compound, including, for example, polymorphs,pseudopolymorphs, solvates, hydrates, unsolvated polymorphs (includinganhydrates), and conformational polymorphs, as well as mixtures thereof,unless a particular crystalline form is referred to.

“Solvate” refers to a crystalline phase of a compound in physicalassociation with one or more molecules of a solvent. The crystallinephase of a compound in physical association with one or more moleculesof water is referred to as a “hydrate.”

“Amorphous form” refers to a form of a compound, or a salt or molecularcomplex of a compound, that lacks long range crystalline order where thex-ray diffraction pattern lacks Bragg reflections.

Crystalline Forms

In an embodiment, the invention provides a crystalline solid form of(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide(Formula (1)). Formula (1) has the following chemical structure:

In an embodiment, the invention provides a composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by an X-ray powder diffraction patterncomprising peaks at 6.4, 8.6, 10.5, 11.6, and 15.7 °2θ±0.2 °2θ. In anembodiment, the composition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by an X-ray powder diffraction patterncomprising peaks at 6.4, 8.6, 10.5, 11.6, and 15.7 °2θ±0.2 °2θ andfurther comprising peaks at 10.9, 12.7, 13.4, 14.3, 14.9, and 18.2°2θ±0.2 °2θ. In another embodiment, the composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by an X-ray powder diffraction patterncomprising peaks selected from the group consisting of 6.4, 8.6, 10.5,10.9, 11.3, 11.6, 12.7, 13.4, 14.3, 14.9, 15.1, 15.7, 16.1, 17.3, 18.2,19.2, 19.39, 19.8, 20.7, 21.1, 21.4, 21.6, 21.9, 22.6, 23.3, 23.6, 24.9,25.2, 25.4, 25.7, 26.1, 26.4, 26.8, 26.9, 27.7, 28.6, 29.1, 29.4, 30.1,30.5, 31.7, 31.9, 32.2, 32.6, 33.1, 33.4, 34.5, 35.9, 36.1, 36.8, 37.4,38.1, 38.9, 39.5, and any combination thereof, with peak positionsmeasured in °2θ±0.2 °2θ. In yet another embodiment, the compositioncomprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by a transmission X-ray powder diffractionpattern substantially the same as the representative X-ray powderdiffraction pattern shown in FIG. 2. In an embodiment, the X-ray powderdiffraction pattern of any of the foregoing embodiments is measured intransmission mode. In an embodiment, the X-ray powder diffractionpattern of any of the foregoing embodiments is measured in reflectionmode.

It is known in the art that an X-ray powder diffraction pattern may beobtained which has one or more measurement errors depending onmeasurement conditions (such as equipment, sample preparation orinstrument used). In particular, it is generally known that intensitiesin an X-ray powder diffraction pattern may vary depending on measurementconditions and sample preparation. For example, persons skilled in theart of X-ray powder diffraction will realise that the relativeintensities of peaks may vary according to the orientation of the sampleunder test and based on the type and settings of the instrument used.The skilled person will also realise that the position of reflectionscan be affected by the precise height at which the sample sits in thediffractometer, the sample's surface planarity, and the zero calibrationof the diffractometer. Hence a person skilled in the art will appreciatethat the diffraction pattern data presented herein is not to beconstrued as absolute and any crystalline form that provides a powerdiffraction pattern substantially the same as those disclosed hereinfall within the scope of the present disclosure. For furtherinformation, see Jenkins and Snyder, Introduction to X-Ray PowderDiffractometry, John Wiley & Sons, 1996.

In each of the foregoing embodiments, composition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is further characterized by at least one of: (1) a Ramanspectrum with at least five peaks selected from the group consisting of1680, 1620, 1609, 1574, 1547, 1514, 1495, 1454, 1433, 1351, 1312, 1255,1232, 1187, 1046, 995, 706, 406, 280, and any combination thereof, withpeak positions measured in cm⁻¹±4 cm⁻¹; (2) an IR spectrum with at leastfive peaks selected from the group consisting of 3367, 3089, 2246, 1682,1621, 1608, 1574, 1514, 1504, 1454, 1428, 1403, 1345, 1303, 1248, 1194,1177, 1149, 1109, 1049, 1023, 1003, 947, 900, 858, 842, 816, 764, 734,729, 701, 689, 665, 623, 612, and any combination thereof, with peakpositions measured in cm⁻¹±4 cm⁻¹; and (3) the absence of water in thecrystal structure.

It is also known in the art that IR and Raman spectra may be obtainedwhich may vary depending on measurement conditions. The instrument,sampling mode (e.g., attenuated total reflectance IR sampling versustransmission IR sampling), and the calibration of the instrument mayaffect the peak positions and intensities. A person skilled in the artwill appreciate that the spectra presented herein is not to be construedas absolute and any crystalline form that provides a spectrumsubstantially the same as those disclosed herein fall within the scopeof the present disclosure. For further information, see Colthup, et al.,Introduction to Infrared and Raman Spectroscopy, 3rd Ed., AcademicPress, 1990.

In an embodiment, the invention provides a composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by at least one of: (1) an X-ray powderdiffraction pattern with at least five peaks selected from the groupconsisting of 6.6, 9.9, 11.0, 13.6, 14.0, 14.3, 18.1, 18.4, 18.9, 19.3,20.2, 21.1, 22.0, 22.2, 22.5, 22.7, 22.9, 23.4, 23.5, 23.9, 24.2, 24.6,25.0, 26.1, 26.6, 26.9, 27.5, 28.2, 31.0, 32.1, 32.4, 32.7, 33.4, 33.9,34.4, and any combination thereof, with peak positions measured in°2θ±0.2 °2θ; (2) a Raman spectrum with at least five peaks selected fromthe group consisting of 1668, 1611, 1580, 1564, 1537, 1506, 1493, 1454,1436, 1416, 1401, 1349, 1321, 1287, 1272, 1252, 1244, 1183, 1165, 1097,1039, 1025, 996, 950, 871, 853, 776, 730, 645, 633, 375, 352, 279, and247 and any combination thereof, with peak positions measured in cm⁻¹±4cm⁻¹; (3) an IR spectrum with at least five peaks selected from thegroup consisting of 3212, 2206, 1665, 1618, 1577, 1548, 1535, 1504,1465, 1452, 1432, 1416, 1397, 1348, 1316, 1243, 1208, 1181, 1164, 1149,1095, 1038, 1004, 948, 891, 869, 821, 776, 736, 716, 643, 617, and anycombination thereof, with peak positions measured in cm⁻¹±4 cm⁻¹; and(4) the presence of water in the crystal structure with a stoichiometryrelative to Formula (1) that is approximately equivalent to atrihydrate.

In an embodiment, the invention provides a composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by at least one of: (1) an X-ray powderdiffraction pattern with at least five peaks selected from the groupconsisting of 10.4, 12.6, 12.8, 17.9, 21.3, 21.7, 23.1, 24.2, 25.2,27.0, and any combination thereof, with at least five peak positionsmeasured in °2θ±0.2 °2θ; (2) an IR spectrum with at least five peaksselected from the group consisting of 3446, 2248, 1667, 1592, 1531,1504, 1428, 1349, 1305, 1243, 1189, 1158, 1089, 1001, 896, 862, 829,780, 759, 736, 699, and any combination thereof, with peak positionsmeasured in cm⁻¹±4 cm⁻¹; and (3) the presence of water in the crystalstructure with a stoichiometry relative to Formula (1) that isapproximately equivalent to a dihydrate.

In an embodiment, the invention provides a composition comprisingcrystalline(5)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefumarate. In an embodiment, the invention provides a compositioncomprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefumarate, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefumarate is characterized by at least one of: (1) an X-ray powderdiffraction pattern with at least five peaks selected from the groupconsisting of 4.9, 5.4, 7.0, 9.8, 10.8, 11.5, 12.1, 14.1, 16.1, 16.6,17.8, 18.5, 19.4, 20.3, 20.5, 21.9, 22.1, 22.5, 23.1, 24.0, 24.8, 26.6,26.8, 27.3, 28.2, and any combination thereof, with peak positionsmeasured in °2θ±0.2 °2θ; and (2) the presence of water in the crystalstructure with a stoichiometry relative to Formula (1) that isapproximately equivalent to a sesquihydrate. In an embodiment, theinvention provides a composition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefumarate Form A, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefumarate Form A is characterized by an X-ray powder diffraction patterncomprising peaks at 4.9, 5.4, 7.0, 10.8, and 11.5 °2θ±0.2 °2θ. In anembodiment, the invention provides a composition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefumarate Form A, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefumarate Form A is characterized by an X-ray powder diffraction patternsubstantially in agreement with an X-ray powder diffraction pattern ofFIG. 41.

In an embodiment, the invention provides a composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidemaleate. In an embodiment, the invention provides a compositioncomprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidemaleate, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidemaleate is characterized by at least one of: (1) an X-ray powderdiffraction pattern with at least five peaks selected from the groupconsisting of 5.3, 9.8, 10.6, 11.6, 13.5, 13.8, 13.9, 14.3, 15.3, 15.6,15.8, 15.9, 16.6, 17.4, 17.5, 18.7, 19.3, 19.6, 19.8, 20.0, 20.9, 21.3,22.1, 22.3, 22.7, 23.2, 23.4, 23.7, 23.9, 24.5, 24.8, 25.2, 25.6, 26.1,26.4, 26.7, 26.9, 27.1, 27.6, 28.9, 29.5, 30.0, 30.3, 30.9, 31.5, 31.9,32.5, 33.9, 35.1, and any combination thereof, with peak positionsmeasured in °2θ±0.2 °2θ; and (2) the presence of water in the crystalstructure with a stoichiometry relative to Formula (1) that isapproximately equivalent to a monohydrate. In an embodiment, theinvention provides a composition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidemaleate Form A, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidemaleate Form A is characterized by an X-ray powder diffraction patterncomprising peaks at 5.3, 9.8, 10.6, 11.6, and 19.3 °2θ±0.2 °2θ. In anembodiment, the invention provides a composition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidemaleate Form A, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidemaleate Form A is characterized by an X-ray powder diffraction patternsubstantially in agreement with the X-ray powder diffraction pattern ofFIG. 42. In an embodiment, the X-ray powder diffraction pattern of anyof the foregoing embodiments is measured in transmission mode. In anembodiment, the X-ray powder diffraction pattern of any of the foregoingembodiments is measured in reflection mode.

In an embodiment, the invention provides a composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidephosphate. In an embodiment, the invention provides a compositioncomprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidephosphate, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidephosphate is characterized by at least one of: (1) an X-ray powderdiffraction pattern with at least five peaks selected from the groupconsisting of 4.5, 6.0, 7.2, 10.4, 12.0, 12.5, 13.1, 14.3, 15.5, 17.4,18.0, 18.3, 18.9, 19.3, 20.2, 20.56, 20.9, 21.4, 21.9, 22.0, 22.6, 22.9,23.1, 23.3, 24.2, 24.6, 25.0, 25.7, 26.2, 26.4, 26.9, 27.3, 27.5, 29.3,30.0, 30.3, 30.5, 30.9, 31.2, 31.9, 35.7, and any combination thereof,with peak positions measured in °2θ±0.2 °2θ; and (2) the presence ofwater in the crystal structure with a stoichiometry relative to Formula(1) that is approximately equivalent to a dihydrate. In an embodiment,the X-ray powder diffraction pattern of any of the foregoing embodimentsis measured in transmission mode. In an embodiment, the X-ray powderdiffraction pattern of any of the foregoing embodiments is measured inreflection mode. In an embodiment, the invention provides a compositioncomprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidephosphate Form A, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidephosphate Form A is characterized by an X-ray powder diffraction patterncomprising peaks at 4.5, 6.0, 10.4, 12.0, and 14.3 °2θ±0.2 °2θ. In anembodiment, the invention provides a composition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidephosphate Form A, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidephosphate Form A is characterized by an X-ray powder diffraction patternsubstantially in agreement with the X-ray powder diffraction pattern ofFIG. 43. In an embodiment, the X-ray powder diffraction pattern of anyof the foregoing embodiments is measured in transmission mode. In anembodiment, the X-ray powder diffraction pattern of any of the foregoingembodiments is measured in reflection mode.

In an embodiment, the invention provides a composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideL-tartrate. In an embodiment, the invention provides a compositioncomprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideL-tartrate, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideL-tartrate is characterized by at least one of: (1) an X-ray powderdiffraction pattern with at least five peaks selected from the groupconsisting of 4.6, 5.5, 7.2, 9.3, 10.7, 10.9, 11.8, 14.3, 14.9, 16.4,17.0, 17.7, 19.2, 19.4, 19.5, 20.3, 21.6, 22.4, 23.3, 23.8, 24.3, 24.5,24.7, 25.1, 25.6, 26.8, 27.2, 27.8, 28.4, 28.7, 29.0, 29.5, 30.0, 30.9,31.6, 32.1, 32.4, 33.0, 33.5, 33.9, and any combination thereof, withpeak positions measured in °2θ±0.2 °2θ; and (2) the presence of water inthe crystal structure with a stoichiometry relative to Formula (1) thatis approximately equivalent to a sesquihydrate. In an embodiment, theinvention provides a composition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideL-tartrate Form A, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideL-tartrate Form A is characterized by an X-ray powder diffractionpattern comprising peaks at 4.6, 5.5, 10.9, 11.8, and 14.9 °2θ±0.2 °2θ.In an embodiment, the invention provides a composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideL-tartrate Form A, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideL-tartrate Form A is characterized by an X-ray powder diffractionpattern substantially in agreement with the X-ray powder diffractionpattern of FIG. 44. In an embodiment, the X-ray powder diffractionpattern of any of the foregoing embodiments is measured in transmissionmode. In an embodiment, the X-ray powder diffraction pattern of any ofthe foregoing embodiments is measured in reflection mode.

In an embodiment, the invention provides a composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidecitrate. In an embodiment, the invention provides a compositioncomprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidecitrate, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidecitrate is characterized by at least one of: (a) an X-ray powderdiffraction pattern with at least five peaks selected from the groupconsisting of 6.1, 6.6, 7.2, 7.9, 8.3, 9.7, 10.8, 11.1, 12.2, 13.5,14.1, 14.9, 15.9, 16.6, 17.5, 17.9, 18.3, 18.9, 19.5, 20.3, 21.5, 21.9,22.7, 23.8, 24.3, 24.8, 26.1, 26.3, 27.2, 27.4, 27.9, 29.3, and anycombination thereof, with peak positions measured in °2θ±0.2 °2θ; (b) aRaman spectrum with at least five peaks selected from the groupconsisting of 3068, 2921, 2237, 1682, 1612, 1551, 1505, 1436, 1332,1313, 1241, 1188, 993, 712, and any combination thereof, with peakpositions measured in cm⁻¹±4 cm⁻¹; (c) an IR spectrum with at least fivepeaks selected from the group consisting 3396, 2234, 1673, 1606, 1537,1428, 1304, 1264, 1200, 1092, 1008, 893, 866, 773, 735, and 693, and anycombination thereof, with peak positions measured in cm⁻¹±4 cm⁻¹; and(d) the presence of water in the crystal structure at a concentrationbetween about 0% to 8% by weight. In an embodiment, the inventionprovides a composition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidecitrate, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidecitrate is characterized by at least one of: (a) an X-ray powderdiffraction pattern with at least five peaks selected from the groupconsisting of 6.1, 6.4, 7.2, 7.9, 8.2, 9.6, 10.9, 12.0, 13.4, 13.8,14.0, 14.9, 15.5, 15.9, 16.4, 17.3, 17.5, 18.2, 18.6, 19.3, 20.1, 20.4,21.4, 21.6, 22.6, 23.2, 23.7, 24.3, 26.0, 27.0, 27.3, 27.8, 29.2, andany combination thereof, with peak positions measured in °2θ±0.2 °2θ;(b) a Raman spectrum with at least five peaks selected from the groupconsisting of 3055, 2920, 2237, 1685, 1612, 1549, 1504, 1436, 1333,1313, 1286, 1240, 1187, 993, 712, and any combination thereof, with peakpositions measured in cm⁻¹±4 cm⁻¹; (c) an IR spectrum with at least fivepeaks selected from the group consisting 3403, 2960, 2872, 2233, 1678,1608, 1582, 1538, 1434, 1403, 1352, 1302, 1253, 1201, 1094, 1055, 1010,967, 895, 813, 772, 750, 735, 693, 612, and any combination thereof,with peak positions measured in cm⁻¹±4 cm⁻¹; and (d) the presence ofwater in the crystal structure at a concentration between about 0% toabout 8% by weight. In an embodiment, the invention provides acomposition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidecitrate Form A, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidecitrate Form A is characterized by an X-ray powder diffraction patterncomprising peaks at 6.1, 7.2, 9.7, 11.1, and 12.2 °2θ±0.2 °2θ. In anembodiment, the invention provides a composition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidecitrate Form A, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidecitrate Form A is characterized by an X-ray powder diffraction patternsubstantially in agreement with the X-ray powder diffraction pattern ofFIG. 58. In an embodiment, the invention provides a compositioncomprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidecitrate Form A, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidecitrate Form A is characterized by an X-ray powder diffraction patterncomprising peaks at 6.1, 7.2, 9.6, 10.9, and 12.0 °2θ±0.2 °2θ. In anembodiment, the X-ray powder diffraction pattern of any of the foregoingembodiments is measured in transmission mode. In an embodiment, theX-ray powder diffraction pattern of any of the foregoing embodiments ismeasured in reflection mode.

In an embodiment, the invention provides a composition comprisingcrystalline(5)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidegentisate. In an embodiment, the invention provides a compositioncomprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidegentisate, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidegentisate is characterized by at least one of: (a) an X-ray powderdiffraction pattern with at least five peaks selected from the groupconsisting of 4.6, 8.2, 9.0, 9.7, 11.8, 12.9, 13.8, 14.5, 15.5, 16.6,16.8, 18.4, 19.6, 20.5, 21.1, 24.1, 24.5, 25.5, 25.8, 26.0, 26.6, 26.9,27.4, 29.8, and any combination thereof, with peak positions measured in°2θ±0.2 °2θ; (b) a Raman spectrum with at least five peaks selected fromthe group consisting of 3057, 2919, 2223, 1681, 1613, 1576, 1552, 1518,1437, 1333, 1312, 1228, 1192, 1156, 990, 716, 485, 257, and anycombination thereof, with peak positions measured in cm⁻¹±4 cm⁻¹; (c) anIR spectrum with at least five peaks selected from the group consisting2957, 1682, 1668, 1602, 1574, 1523, 1504, 1481, 1429, 1377, 1346, 1302,1274, 1228, 1157, 1092, 1010, 939, 896, 865, 826, 810, 778, 748, 734,686, 660, 617, and any combination thereof, with peak positions measuredin cm⁻¹±4 cm⁻¹; and (d) the presence of water in the crystal structurewith a stoichiometry relative to Formula (1) that is approximatelyequivalent to a monohydrate. In an embodiment, the invention provides acomposition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidegentisate Form A, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidegentisate Form A is characterized by an X-ray powder diffraction patterncomprising peaks at 4.6, 9.0, 12.9, 13.8, and 19.6 °2θ±0.2 °2θ. In anembodiment, the invention provides a composition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidegentisate Form A, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidegentisate Form A is characterized by an X-ray powder diffraction patternsubstantially in agreement with the X-ray powder diffraction pattern ofFIG. 60. In an embodiment, the X-ray powder diffraction pattern of anyof the foregoing embodiments is measured in transmission mode. In anembodiment, the X-ray powder diffraction pattern of any of the foregoingembodiments is measured in reflection mode.

In an embodiment, the invention provides a composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideoxalate. In an embodiment, the invention provides a compositioncomprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideoxalate, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideoxalate is characterized by at least one of: (a) an X-ray powderdiffraction pattern with at least five peaks selected from the groupconsisting of 5.5, 5.8, 7.4, 9.3, 11.0, 11.5, 12.7, 15.2, 16.5, 17.3,18.5, 18.7, 19.1, 19.7, 20.2, 20.8, 22.0, 22.3, 23.3, 23.6, 24.8, 27.4,28.6, 29.3, 29.6, 31.2, 33.1, and any combination thereof, with peakpositions measured in °2θ±0.2 °2θ; (b) a Raman spectrum with at leastfive peaks selected from the group consisting of 3073, 2992, 2950, 2922,2247, 1671, 1612, 1584, 1552, 1504, 1469, 1440, 1336, 1311, 1273, 1235,1191, 1162, 1095, 1012, 897, 718, 633, 409, 370, 263, and anycombination thereof, with peak positions measured in cm⁻¹±4 cm⁻¹; (c) anIR spectrum with at least five peaks selected from the group consisting3419, 2249, 1670, 1615, 1544, 1503, 1438, 1391, 1334, 1304, 1262, 1195,1151, 1126, 1093, 1013, 894, 877, 823, 783, 765, 738, 652, and anycombination thereof, with peak positions measured in cm⁻¹±4 cm⁻¹; and(d) the presence of water in the crystal structure at a concentrationbetween about 0% to about 9% by weight. In an embodiment, the inventionprovides a composition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideoxalate Form A, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideoxalate Form A is characterized by an X-ray powder diffraction patterncomprising peaks at 5.5, 5.8, 9.3, 11.5, and 12.7 °2θ±0.2 °2θ. In anembodiment, the invention provides a composition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideoxalate Form A, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideoxalate Form A is characterized by an X-ray powder diffraction patternsubstantially in agreement with the X-ray powder diffraction pattern ofFIG. 61. In an embodiment, the X-ray powder diffraction pattern of anyof the foregoing embodiments is measured in transmission mode. In anembodiment, the X-ray powder diffraction pattern of any of the foregoingembodiments is measured in reflection mode.

In an embodiment, the invention provides a composition comprisingcrystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidesulfate. In an embodiment, the invention provides a compositioncomprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidesulfate, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidesulfate is characterized by at least one of: (a) an X-ray powderdiffraction pattern with at least five peaks selected from the groupconsisting of 4.6, 5.0, 8.0, 9.0, 9.8, 12.0, 12.7, 13.2, 14.6, 15.0,15.6, 16.2, 17.5, 17.9, 19.8, 20.2, 21.9, 23.8, 24.4, 24.9, 25.7, 26.0,27.2, 29.5, 30.4, 31.6, 32.5, and any combination thereof, with peakpositions measured in °2θ±0.2 °2θ; (b) a Raman spectrum with at leastfive peaks selected from the group consisting of 3115, 2977, 2926, 2224,1675, 1611, 1537, 1498, 1449, 1409, 1361, 1327, 1310, 1288, 1243, 1198,1155, 1042, 1009, 978, 948, 906, 849, 771, 713, 652, 632, 464, 370, 254,and any combination thereof, with peak positions measured in cm⁻¹±4cm⁻¹; (c) an IR spectrum with at least five peaks selected from thegroup consisting 3430, 3101, 3029, 2225, 1667, 1633, 1615, 1598, 1563,1557, 1508, 1428, 1350, 1328, 1308, 1276, 1225, 1088, 1036, 1018, 925,891, 848, 816, 783, 736, 723, 694, 612, and any combination thereof,with peak positions measured in cm⁻¹±4 cm⁻¹; and (d) the presence ofwater in the crystal structure at a concentration between about 2.5% toabout 12.5% by weight. In an embodiment, the invention provides acomposition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidesulfate Form A, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidesulfate Form A is characterized by an X-ray powder diffraction patterncomprising peaks at 4.6, 9.0, 9.8, 17.5, and 18.0 °2θ±0.2 °2θ. In anembodiment, the invention provides a composition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidesulfate Form A, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidesulfate Form A is characterized by an X-ray powder diffraction patternsubstantially in agreement with the X-ray powder diffraction pattern ofFIG. 66. In an embodiment, the X-ray powder diffraction pattern of anyof the foregoing embodiments is measured in transmission mode. In anembodiment, the X-ray powder diffraction pattern of any of the foregoingembodiments is measured in reflection mode.

In an embodiment, the invention provides a composition comprising acocrystal of(5)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideand L-arabitol. In an embodiment, the invention provides a compositioncomprising a cocrystal of(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideand L-arabitol, wherein the cocrystal of(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideL-arabitol is characterized by at least one of: (a) an X-ray powderdiffraction pattern with at least five peaks selected from the groupconsisting of 5.6, 7.0, 9.2, 11.2, 12.9, 13.5, 14.0, 14.8, 15.3, 16.8,17.9, 18.2, 18.4, 21.1, 21.4, 22.5, 22.9, 23.9, 24.8, 25.5, 26.0, 26.6,27.7, 28.2, 28.8, 29.4, and any combination thereof, with peak positionsmeasured in °2θ±0.2 °2θ; (b) a Raman spectrum with at least five peaksselected from the group consisting of 2917, 2241, 1674, 1610, 1581,1565, 1529, 1494, 1455, 1348, 1325, 1309, 1264, 1242, 1189, 1164, 999,872, 279, and any combination thereof, with peak positions measured incm⁻¹±4 cm⁻¹; (c) an IR spectrum with at least five peaks selected fromthe group consisting 3471, 3188, 2924, 2239, 1670, 1637, 1621, 1603,1579, 1524, 1505, 1435, 1346, 1306, 1274, 1242, 1203, 1135, 1090, 1049,1009, 998, 950, 902, 892, 862, 821, 783, 739, 726, 711, 694, 637, 621,and any combination thereof, with peak positions measured in cm⁻¹±4cm⁻¹; and (d) the presence of water in the crystal structure at aconcentration between about 0% to about 5% by weight. In an embodiment,the X-ray powder diffraction pattern of any of the foregoing embodimentsis measured in transmission mode. In an embodiment, the X-ray powderdiffraction pattern of any of the foregoing embodiments is measured inreflection mode.

In an embodiment, the invention provides a composition comprising acocrystal of(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideand L-proline. In an embodiment, the invention provides a compositioncomprising a cocrystal of(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideand L-proline, wherein the cocrystal of(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideL-proline is characterized by at least one of: (a) an X-ray powderdiffraction pattern with at least five peaks selected from the groupconsisting of 7.0, 10.5, 14.0, 14.8, 17.1, 17.6, 21.1, 22.1, and anycombination thereof, with peak positions measured in °2θ±0.2 °2θ; (b) aRaman spectrum with at least five peaks selected from the groupconsisting of 3064, 2971, 2919, 2881, 2242, 1675, 1607, 1577, 1530,1484, 1454, 1434, 1344, 1323, 1311, 1264, 1243, 1185, 1157, 1035, 1020,993, 370, 275, 190, 153, and any combination thereof, with peakpositions measured in cm⁻¹±4 cm⁻¹; (c) an IR spectrum with at least fivepeaks selected from the group consisting 3471, 3310, 3108, 2358, 1672,1615, 1526, 1495, 1452, 1431, 1399, 1342, 1305, 1262, 1241, 1184, 1148,1091, 1038, 1014, 991, 944, 890, 872, 837, 816, 775, 756, 737, 713, 668,and any combination thereof, with peak positions measured in cm⁻¹±4cm⁻¹; and (d) the absence of water in the crystal structure. In anembodiment, the X-ray powder diffraction pattern of any of the foregoingembodiments is measured in transmission mode. In an embodiment, theX-ray powder diffraction pattern of any of the foregoing embodiments ismeasured in reflection mode.

In an embodiment, the invention provides a composition comprising acocrystal of(5)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideand D-sorbitol. In an embodiment, the invention provides a compositioncomprising a cocrystal of(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideand D-sorbitol, wherein the cocrystal of(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideD-sorbitol is characterized by at least one of: (a) an X-ray powderdiffraction pattern with at least five peaks selected from the groupconsisting of 3.0, 4.4, 5.1, 6.0, 7.3, 8.3, 8.6, 10.1, 12.3, 12.7, 13.4,13.0, 14.3, 14.5, 15.6, 16.5, 17.1, 18.0, 18.5, 18.9, 19.4, 19.8, 20.5,21.2, 21.8, 22.3, 22.6, 24.4, 25.5, 27.3, 28.0, 32.9, and anycombination thereof, with peak positions measured in °2θ±0.2 °2θ; (b) aRaman spectrum with at least five peaks selected from the groupconsisting of 3071, 2921, 2246, 1682, 1610, 1579, 1531, 1493, 1453,1437, 1343, 1311, 1246, 1183, 1162, 1039, 1000, 950, 646, and anycombination thereof, with peak positions measured in cm⁻¹±4 cm⁻¹; (c) anIR spectrum with at least five peaks selected from the group consisting3207, 2244, 1681, 1666, 1651, 1615, 1578, 1548, 1531, 1504, 1463, 1434,1418, 1398, 1311, 1243, 1207, 1149, 1112, 1093, 1052, 1017, 1004, 948,891, 867, 821, 777, 735, 724, 707, 643, 618, and any combinationthereof, with peak positions measured in cm⁻¹±4 cm⁻¹; and (d) thepresence of water in the crystal structure at a concentration betweenabout 0% to about 13% by weight. In an embodiment, the X-ray powderdiffraction pattern of any of the foregoing embodiments is measured intransmission mode. In an embodiment, the X-ray powder diffractionpattern of any of the foregoing embodiments is measured in reflectionmode.

In an embodiment, the invention provides a composition comprising acocrystal of(5)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideand succinic acid. In an embodiment, the invention provides acomposition comprising a cocrystal of(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideand succinic acid, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidesuccinic acid is characterized by at least one of: (a) an X-ray powderdiffraction pattern with at least five peaks selected from the groupconsisting of 5.2, 7.5, 8.0, 8.4, 10.3, 12.1, 13.3, 14.2, 14.7, 15.5,16.0, 17.1, 17.5, 18.1, 18.6, 19.2, 19.6, 20.5, 21.2, 22.5, 22.8, 23.2,23.6, 25.5, 26.2, 27.4, 28.2, 28.7, 29.4, 30.3, 31.0, 31.6, 32.2, 33.2,35.5, and any combination thereof, with peak positions measured in°2θ±0.2 °2θ; (b) a Raman spectrum with at least five peaks selected fromthe group consisting of 2973, 2922, 2252, 1670, 1613, 1580, 1566, 1545,1529, 1496, 1450, 1347, 1330, 1307, 1270, 1244, 1190, 1160, 1036, 1010,844, 728, 634, 411, 237, 200, 138, and any combination thereof, withpeak positions measured in cm⁻¹±4 cm⁻¹; (c) an IR spectrum with at leastfive peaks selected from the group consisting 2359, 1712, 1668, 1620,1603, 1578, 1526, 1494, 1432, 1417, 1403, 1367, 1346, 1304, 1246, 1220,1174, 1162, 1130, 1096, 1035, 1009, 993, 965, 950, 896, 863, 851, 838,778, 754, 734, 726, 712, 672, 636, 624, 606, and any combinationthereof, with peak positions measured in cm⁻¹±4 cm⁻¹. In an embodiment,the X-ray powder diffraction pattern of any of the foregoing embodimentsis measured in transmission mode. In an embodiment, the X-ray powderdiffraction pattern of any of the foregoing embodiments is measured inreflection mode.

Amorphous Forms

In an embodiment, the invention provides a composition comprising anamorphous solid dispersion of(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide(Formula (1)) and a polymer, wherein the polymer is selected from thegroup consisting of hydroxypropylmethylcellulose acetate succinate,polyvinylpyrrolidone, vinylpyrrolidone-vinyl acetate copolymer,methacrylic acid-ethyl acrylate copolymer, methacrylic acid-methylmethacrylate copolymer, and a mixture of vinylpyrrolidone-vinyl acetatecopolymer and hydroxypropylmethylcellulose, wherein the concentration ofthe polymer is between about 20% and about 95% by weight of thecomposition, and wherein the(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideand polymer are molecularly dispersed in each other. Such a compositionmay be referred to as an amorphous solid dispersion of Formula (1). Inan embodiment, the concentration of the polymer is selected from thegroup consisting of about 30% by weight of the composition, about 35% byweight of the composition, about 40% by weight of the composition, about45% by weight of the composition, about 50% by weight of thecomposition, about 55% by weight of the composition, about 60% by weightof the composition, about 65% by weight of the composition, about 70% byweight of the composition, about 75% by weight of the composition, about80% by weight of the composition, about 85% by weight of thecomposition, about 90% by weight of the composition, and about 95% byweight of the composition. In an embodiment, the concentration of thepolymer is between about 30% and about 95% by weight of the composition.In an embodiment, the concentration of the polymer is between about 40%and about 95% by weight of the composition. In an embodiment, theconcentration of the polymer is between about 50% and about 95% byweight of the composition. In an embodiment, the concentration of thepolymer is between about 60% and about 95% by weight of the composition.In an embodiment, the concentration of the polymer is between about 70%and about 95% by weight of the composition. In an embodiment, theconcentration of the polymer is between about 80% and about 95% byweight of the composition. In any of the foregoing embodiments, theamorphous solid dispersion is an amorphous solid molecular dispersion.In any of the foregoing embodiments, the amorphous solid dispersion ischaracterized by a single glass transition temperature (T_(g)) whenanalyzed by differential scanning calorimetry, including modulateddifferential scanning calorimetry or temperature-modulated differentialscanning calorimetry. In any of the foregoing embodiments, the amorphoussolid dispersion is characterized by domains containing highconcentrations of Formula (1) that are of the order of 30 nm or less inapproximate size. In any of the foregoing embodiments, the amorphoussolid dispersion is an intimate mixture of Formula (1) and a polymer.

In an embodiment, an amorphous solid dispersion of Formula (1) furthercomprises an extragranular acidulant. In an embodiment, an amorphoussolid dispersion of Formula (1) further comprises an extragranularacidulant, wherein the extragranular acidulant is selected from thegroup consisting of fumaric acid, succinic acid, D-tartaric acid,L-tartaric acid, racemic tartaric acid, ascorbic acid, isoascorbic acid(also known as erythorbic acid and D-araboascorbic acid), alginic acid,Protacid F 120 NM, Protacid AR 1112 (also known as Kelacid NF), Carbopol971P (carboxypolymethylene), and Carbomer 941 (polyacrylic acid), andcombinations thereof. In an embodiment, an amorphous solid dispersion ofFormula (1) further comprises an extragranular acidulant, wherein theextragranular acidulant is fumaric acid at a concentration of betweenabout 15% to about 33% by weight. In an embodiment, an amorphous soliddispersion of Formula (1) further comprises an extragranular acidulant,wherein the extragranular acidulant is alginic acid at a concentrationof between about 5% to about 33% by weight. In an embodiment, anamorphous solid dispersion of Formula (1) further comprises anextragranular acidulant, wherein the extragranular acidulant isL-tartaric acid at a concentration of between about 25% to about 33% byweight. In an embodiment, an amorphous solid dispersion of Formula (1)further comprises an extragranular acidulant, wherein the extragranularacidulant is ascorbic acid at a concentration of between about 20% toabout 50% by weight and Carbomer 941 (polyacrylic acid) at aconcentration of between about 2.5% to about 10% by weight. In anembodiment, an amorphous solid dispersion of Formula (1) furthercomprises an extragranular acidulant, wherein the extragranularacidulant is ascorbic acid at a concentration of between about 20% toabout 50% by weight and Carbopol 971P (carboxypolymethylene) at aconcentration of between about 2.5% to about 10% by weight. In anembodiment, an amorphous solid dispersion of Formula (1) furthercomprises an extragranular acidulant, wherein the extragranularacidulant is fumaric acid at a concentration of between about 5% toabout 15% by weight and alginic acid at a concentration of about 15% toabout 33% by weight. In an embodiment, an amorphous solid dispersion ofFormula (1) further comprises an extragranular acidulant, wherein theextragranular acidulant is L-tartaric acid at a concentration of betweenabout 5% to 15% by weight and alginic acid at a concentration of betweenabout 15% to about 33% by weight.

In an embodiment, the invention provides for preparation of any of theforegoing amorphous solid dispersions comprising a step selected fromthe group consisting of spray drying, hot melt extrusion,lyophilization, co-grinding, co-milling, evaporation, and combinationsthereof. Suitable methods for preparation of any of the foregoingamorphous solid dispersions may also be found in U.S. Pat. Nos.6,548,555; 8,173,142; 8,236,328; and 8,263,128, the disclosures of whichare incorporated by reference herein.

Pharmaceutical Compositions

In an embodiment, the invention provides a pharmaceutical compositioncomprising a crystalline polymorphic form of the free base of the BTKinhibitor of Formula (1). In an embodiment, the invention provides apharmaceutical composition comprising a crystalline solvate of the freebase of Formula (1). In an embodiment, the invention provides apharmaceutical composition comprising a crystalline hydrate of the freebase of the BTK inhibitor of Formula (1). In an embodiment, theinvention provides a pharmaceutical composition comprising a crystallinesalt of Formula (1). In an embodiment, the invention provides apharmaceutical composition comprising an amorphous form of the BTKinhibitor of Formula (1).

The pharmaceutical compositions are typically formulated to provide atherapeutically effective amount o of a solid form of the BTK inhibitorof Formula (1), as the active ingredient, or a pharmaceuticallyacceptable salt, ester, prodrug, solvate, hydrate or derivative thereof.Where desired, the pharmaceutical compositions contain apharmaceutically acceptable salt thereof, and one or morepharmaceutically acceptable excipients, carriers, including inert soliddiluents and fillers, diluents, permeation enhancers, solubilizers, oradjuvants. The pharmaceutical compositions may also contain anacidulant, as described herein, for reducing or overcoming the effectsof acid reducing agents on the exposure of BTK inhibitor of Formula (1).

In some embodiments, the concentration of a solid form of the BTKinhibitor of Formula (1), provided in the pharmaceutical compositions ofthe invention is independently less than, for example, 100%, 90%, 80%,70%, 60%, 50%, 40%, 30%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%,11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, 0.4%, 0.3%, 0.2%,0.1%, 0.09%, 0.08%, 0.07%, 0.06%, 0.05%, 0.04%, 0.03%, 0.02%, 0.01%,0.009%, 0.008%, 0.007%, 0.006%, 0.005%, 0.004%, 0.003%, 0.002%, or0.001%, w/w, w/v, or v/v, relative to the total mass or volume of thepharmaceutical composition.

In some embodiments, the concentration of a solid forms of the BTKinhibitor of Formula (1), provided in the pharmaceutical compositions ofthe invention is independently greater than 90%, 80%, 70%, 60%, 50%,40%, 30%, 20%, 19.75%, 19.50%, 19.25% 19%, 18.75%, 18.50%, 18.25% 18%,17.75%, 17.50%, 17.25% 17%, 16.75%, 16.50%, 16.25% 16%, 15.75%, 15.50%,15.25% 15%, 14.75%, 14.50%, 14.25% 14%, 13.75%, 13.50%, 13.25% 13%,12.75%, 12.50%, 12.25% 12%, 11.75%, 11.50%, 11.25% 11%, 10.75%, 10.50%,10.25% 10%, 9.75%, 9.50%, 9.25% 9%, 8.75%, 8.50%, 8.25% 8%, 7.75%,7.50%, 7.25% 7%, 6.75%, 6.50%, 6.25% 6%, 5.75%, 5.50%, 5.25% 5%, 4.75%,4.50%, 4.25%, 4%, 3.75%, 3.50%, 3.25%, 3%, 2.75%, 2.50%, 2.25%, 2%,1.75%, 1.50%, 125%, 1%, 0.5%, 0.4%, 0.3%, 0.2%, 0.1%, 0.09%, 0.08%,0.07%, 0.06%, 0.05%, 0.04%, 0.03%, 0.02%, 0.01%, 0.009%, 0.008%, 0.007%,0.006%, 0.005%, 0.004%, 0.003%, 0.002%, 0.001%, 0.0009%, 0.0008%,0.0007%, 0.0006%, 0.0005%, 0.0004%, 0.0003%, 0.0002% or 0.0001% w/w,w/v, or v/v, relative to the total mass or volume of the pharmaceuticalcomposition.

In some embodiments, the concentration of a solid form of the BTKinhibitor of Formula (1), of the invention is independently in the rangefrom approximately 0.0001% to approximately 50%, approximately 0.001% toapproximately 40%, approximately 0.01% to approximately 30%,approximately 0.02% to approximately 29%, approximately 0.03% toapproximately 28%, approximately 0.04% to approximately 27%,approximately 0.05% to approximately 26%, approximately 0.06% toapproximately 25%, approximately 0.07% to approximately 24%,approximately 0.08% to approximately 23%, approximately 0.09% toapproximately 22%, approximately 0.1% to approximately 21%,approximately 0.2% to approximately 20%, approximately 0.3% toapproximately 19%, approximately 0.4% to approximately 18%,approximately 0.5% to approximately 17%, approximately 0.6% toapproximately 16%, approximately 0.7% to approximately 15%,approximately 0.8% to approximately 14%, approximately 0.9% toapproximately 12% or approximately 1% to approximately 10% w/w, w/v orv/v, relative to the total mass or volume of the pharmaceuticalcomposition.

In some embodiments, the concentration of a solid form of the BTKinhibitor of Formula (1), of the invention is independently in the rangefrom approximately 0.001% to approximately 10%, approximately 0.01% toapproximately 5%, approximately 0.02% to approximately 4.5%,approximately 0.03% to approximately 4%, approximately 0.04% toapproximately 3.5%, approximately 0.05% to approximately 3%,approximately 0.06% to approximately 2.5%, approximately 0.07% toapproximately 2%, approximately 0.08% to approximately 1.5%,approximately 0.09% to approximately 1%, approximately 0.1% toapproximately 0.9% w/w, w/v, or v/v, relative to the total mass orvolume of the pharmaceutical composition.

In some embodiments, the amount of a solid form of the BTK inhibitor ofFormula (1), of the invention is independently equal to or less than 3.0g, 2.5 g, 2.0 g, 1.5 g, 1.0 g, 0.95 g, 0.9 g, 0.85 g, 0.8 g, 0.75 g, 0.7g, 0.65 g, 0.6 g, 0.55 g, 0.5 g, 0.45 g, 0.4 g, 0.35 g, 0.3 g, 0.25 g,0.2 g, 0.15 g, 0.1 g, 0.09 g, 0.08 g, 0.07 g, 0.06 g, 0.05 g, 0.04 g,0.03 g, 0.02 g, 0.01 g, 0.009 g, 0.008 g, 0.007 g, 0.006 g, 0.005 g,0.004 g, 0.003 g, 0.002 g, 0.001 g, 0.0009 g, 0.0008 g, 0.0007 g, 0.0006g, 0.0005 g, 0.0004 g, 0.0003 g, 0.0002 g or 0.0001 g.

In some embodiments, the amount of a solid form of the BTK inhibitor ofFormula (1), of the invention is independently more than 0.0001 g,0.0002 g, 0.0003 g, 0.0004 g, 0.0005 g, 0.0006 g, 0.0007 g, 0.0008 g,0.0009 g, 0.001 g, 0.0015 g, 0.002 g, 0.0025 g, 0.003 g, 0.0035 g, 0.004g, 0.0045 g, 0.005 g, 0.0055 g, 0.006 g, 0.0065 g, 0.007 g, 0.0075 g,0.008 g, 0.0085 g, 0.009 g, 0.0095 g, 0.01 g, 0.015 g, 0.02 g, 0.025 g,0.03 g, 0.035 g, 0.04 g, 0.045 g, 0.05 g, 0.055 g, 0.06 g, 0.065 g, 0.07g, 0.075 g, 0.08 g, 0.085 g, 0.09 g, 0.095 g, 0.1 g, 0.15 g, 0.2 g, 0.25g, 0.3 g, 0.35 g, 0.4 g, 0.45 g, 0.5 g, 0.55 g, 0.6 g, 0.65 g, 0.7 g,0.75 g, 0.8 g, 0.85 g, 0.9 g, 0.95 g, 1 g, 1.5 g, 2 g, 2.5, or 3 g.

Each of the solid forms of the BTK inhibitor of Formula (1), accordingto the invention is effective over a wide dosage range. For example, inthe treatment of adult humans, dosages independently range from 0.01 to1000 mg, from 0.5 to 100 mg, from 1 to 50 mg per day, from 2 to 40 mgper day, and from 5 to 25 mg per day are examples of dosages that may beused. The exact dosage will depend upon the route of administration, theform in which the compound is administered, the gender and age of thesubject to be treated, the body weight of the subject to be treated, andthe preference and experience of the attending physician.

In selected embodiments, the invention provides a pharmaceuticalcomposition for oral administration containing the BTK inhibitor ofFormula (1), and a pharmaceutical excipient suitable for oraladministration.

In selected embodiments, the invention provides a solid pharmaceuticalcomposition for oral administration containing: (i) an effective amountof the BTK inhibitor of Formula (1), and (ii) a pharmaceutical excipientsuitable for oral administration. In selected embodiments, thecomposition further contains (iii) an effective amount of another activepharmaceutical ingredient.

In selected embodiments, the pharmaceutical composition may be a liquidpharmaceutical composition suitable for oral consumption. Pharmaceuticalcompositions of the invention suitable for oral administration can bepresented as discrete dosage forms, such as capsules, sachets, ortablets, or liquids or aerosol sprays each containing a predeterminedamount of an active ingredient as a powder or in granules, a solution,or a suspension in an aqueous or non-aqueous liquid, an oil-in-wateremulsion, or a water-in-oil emulsion. Pharmaceutical compositions of theinvention also include powder for reconstitution, powders for oralconsumptions, bottles (such as powder or liquid in bottle), orallydissolving films, lozenges, pastes, tubes, gums, and packs. Such dosageforms can be prepared by any of the methods of pharmacy, but all methodsinclude the step of bringing the active ingredient(s) into associationwith the carrier, which constitutes one or more necessary ingredients.In general, the compositions are prepared by uniformly and intimatelyadmixing the active ingredient(s) with liquid carriers or finely dividedsolid carriers or both, and then, if necessary, shaping the product intothe desired presentation. For example, a tablet can be prepared bycompression or molding, optionally with one or more accessoryingredients. Compressed tablets can be prepared by compressing in asuitable machine the active ingredient in a free-flowing form such aspowder or granules, optionally mixed with an excipient such as, but notlimited to, a binder, a lubricant, an inert diluent, and/or a surfaceactive or dispersing agent. Molded tablets can be made by molding in asuitable machine a mixture of the powdered compound moistened with aninert liquid diluent.

The invention further encompasses anhydrous pharmaceutical compositionsand dosage forms since water can facilitate the degradation of somecompounds. For example, water may be added (e.g., 5%) in thepharmaceutical arts as a means of simulating long-term storage in orderto determine characteristics such as shelf-life or the stability offormulations over time. Anhydrous pharmaceutical compositions and dosageforms of the invention can be prepared using anhydrous or low moisturecontaining ingredients and low moisture or low humidity conditions.Pharmaceutical compositions and dosage forms of the invention whichcontain lactose can be made anhydrous if substantial contact withmoisture and/or humidity during manufacturing, packaging, and/or storageis expected. An anhydrous pharmaceutical composition may be prepared andstored such that its anhydrous nature is maintained. Accordingly,anhydrous compositions may be packaged using materials known to preventexposure to water such that they can be included in suitable formularykits. Examples of suitable packaging include, but are not limited to,hermetically sealed foils, plastic or the like, unit dose containers,blister packs, and strip packs.

Each of the solid forms of the BTK inhibitor of Formula (1), can becombined in an intimate admixture with a pharmaceutical carrieraccording to conventional pharmaceutical compounding techniques. Thecarrier can take a wide variety of forms depending on the form ofpreparation desired for administration. In preparing the compositionsfor an oral dosage form, any of the usual pharmaceutical media can beemployed as carriers, such as, for example, water, glycols, oils,alcohols, flavoring agents, preservatives, coloring agents, and the likein the case of oral liquid preparations (such as suspensions, solutions,and elixirs) or aerosols; or carriers such as starches, sugars,micro-crystalline cellulose, diluents, granulating agents, lubricants,glidants, binders, and disintegrating agents can be used in the case oforal solid preparations, in some embodiments without employing the useof lactose. For example, suitable carriers include powders, capsules,and tablets, with the solid oral preparations. If desired, tablets canbe coated by standard aqueous or nonaqueous techniques.

Binders suitable for use in pharmaceutical compositions and dosage formsinclude, but are not limited to, corn starch, potato starch, or otherstarches, gelatin, natural and synthetic gums such as acacia, sodiumalginate, alginic acid, other alginates, powdered tragacanth, guar gum,cellulose and its derivatives (e.g., ethyl cellulose, cellulose acetate,carboxymethyl cellulose calcium, sodium carboxymethyl cellulose),polyvinyl pyrrolidone, methyl cellulose, pre-gelatinized starch,hydroxypropyl methyl cellulose, microcrystalline cellulose, and mixturesthereof.

Examples of suitable fillers for use in the pharmaceutical compositionsand dosage forms disclosed herein include, but are not limited to, talc,calcium carbonate (e.g., granules or powder), microcrystallinecellulose, powdered cellulose, dextrates, kaolin, mannitol, silicicacid, sorbitol, starch, pre-gelatinized starch, and mixtures thereof.

Disintegrants may be used in the compositions of the invention toprovide tablets that disintegrate when exposed to an aqueousenvironment. Too much of a disintegrant may produce tablets whichdisintegrate in the bottle. Too little may be insufficient fordisintegration to occur, thus altering the rate and extent of release ofthe active ingredients from the dosage form. Thus, a sufficient amountof disintegrant that is neither too little nor too much to detrimentallyalter the release of the active ingredient(s) may be used to form thedosage forms of the compounds disclosed herein. The amount ofdisintegrant used may vary based upon the type of formulation and modeof administration, and may be readily discernible to those of ordinaryskill in the art. About 0.5 to about 15 weight percent of disintegrant,or about 1 to about 5 weight percent of disintegrant, may be used in thepharmaceutical composition. Disintegrants that can be used to formpharmaceutical compositions and dosage forms of the invention include,but are not limited to, agar-agar, alginic acid, calcium carbonate,microcrystalline cellulose, croscarmellose sodium, crospovidone,polacrilin potassium, sodium starch glycolate, potato or tapioca starch,other starches, pre-gelatinized starch, other starches, clays, otheralgins, other celluloses, gums or mixtures thereof.

Lubricants which can be used to form pharmaceutical compositions anddosage forms of the invention include, but are not limited to, calciumstearate, magnesium stearate, mineral oil, light mineral oil, glycerin,sorbitol, mannitol, polyethylene glycol, other glycols, stearic acid,sodium stearyl fumarate, sodium lauryl sulfate, talc, hydrogenatedvegetable oil (e.g., peanut oil, cottonseed oil, sunflower oil, sesameoil, olive oil, corn oil, and soybean oil), zinc stearate, ethyl oleate,ethylaureate, agar, or mixtures thereof. Additional lubricants include,for example, a SYLOID® silica gel, a coagulated aerosol of syntheticsilica, silicified microcrystalline cellulose, or mixtures thereof. Alubricant can optionally be added, in an amount of less than about 1weight percent of the pharmaceutical composition.

When aqueous suspensions and/or elixirs are desired for oraladministration, the essential active ingredient therein may be combinedwith various sweetening or flavoring agents, coloring matter or dyesand, if so desired, emulsifying and/or suspending agents, together withsuch diluents as water, ethanol, propylene glycol, glycerin and variouscombinations thereof.

The tablets can be uncoated or coated by known techniques to delaydisintegration and absorption in the gastrointestinal tract and therebyprovide a sustained action over a longer period. For example, a timedelay material such as glyceryl monostearate or glyceryl distearate canbe employed. Formulations for oral use can also be presented as hardgelatin capsules wherein the active ingredient is mixed with an inertsolid diluent, for example, calcium carbonate, calcium phosphate orkaolin, or as soft gelatin capsules wherein the active ingredient ismixed with water or an oil medium, for example, peanut oil, liquidparaffin or olive oil.

Surfactants which can be used to form pharmaceutical compositions anddosage forms of the invention include, but are not limited to,hydrophilic surfactants, lipophilic surfactants, and mixtures thereof.That is, a mixture of hydrophilic surfactants may be employed, a mixtureof lipophilic surfactants may be employed, or a mixture of at least onehydrophilic surfactant and at least one lipophilic surfactant may beemployed.

An empirical parameter used to characterize the relative hydrophilicityand hydrophobicity of non-ionic amphiphilic compounds is thehydrophilic-lipophilic balance (“HLB” value). A suitable hydrophilicsurfactant may generally have an HLB value of at least 10, whilesuitable lipophilic surfactants may generally have an HLB value of orless than about 10. Surfactants with lower HLB values are morelipophilic or hydrophobic, and have greater solubility in oils, whilesurfactants with higher HLB values are more hydrophilic, and havegreater solubility in aqueous solutions. Hydrophilic surfactants aregenerally considered to be those compounds having an HLB value greaterthan about 10, as well as anionic, cationic, or zwitterionic compoundsfor which the HLB scale is not generally applicable. Similarly,lipophilic (i.e., hydrophobic) surfactants are compounds having an HLBvalue equal to or less than about 10. However, HLB value of a surfactantis merely a rough guide generally used to enable formulation ofindustrial, pharmaceutical and cosmetic emulsions.

Hydrophilic surfactants may be either ionic or non-ionic. Suitable ionicsurfactants include, but are not limited to, alkylammonium salts;fusidic acid salts; fatty acid derivatives of amino acids,oligopeptides, and polypeptides; glyceride derivatives of amino acids,oligopeptides, and polypeptides; lecithins and hydrogenated lecithins;lysolecithins and hydrogenated lysolecithins; phospholipids andderivatives thereof; lysophospholipids and derivatives thereof;carnitine fatty acid ester salts; salts of alkylsulfates; fatty acidsalts; sodium docusate; acylactylates; mono- and di-acetylated tartaricacid esters of mono- and di-glycerides; succinylated mono- anddi-glycerides; citric acid esters of mono- and di-glycerides; andmixtures thereof.

Within the aforementioned group, ionic surfactants include, by way ofexample: lecithins, lysolecithin, phospholipids, lysophospholipids andderivatives thereof; carnitine fatty acid ester salts; salts ofalkylsulfates; fatty acid salts; sodium docusate; acylactylates; mono-and di-acetylated tartaric acid esters of mono- and di-glycerides;succinylated mono- and di-glycerides; citric acid esters of mono- anddi-glycerides; and mixtures thereof.

Ionic surfactants may be the ionized forms of lecithin, lysolecithin,phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol,phosphatidic acid, phosphatidylserine, lysophosphatidylcholine,lysophosphatidylethanolamine, lysophosphatidylglycerol, lysophosphatidicacid, lysophosphatidylserine, PEG-phosphatidylethanolamine,PVP-phosphatidylethanolamine, lactylic esters of fatty acids,stearoyl-2-lactylate, stearoyllactylate, succinylated monoglycerides,mono/diacetylated tartaric acid esters of mono/diglycerides, citric acidesters of mono/diglycerides, cholylsarcosine, caproate, caprylate,caprate, laurate, myristate, palmitate, oleate, ricinoleate, linoleate,linolenate, stearate, lauryl sulfate, teracecyl sulfate, docusate,lauroyl carnitines, palmitoyl carnitines, myristoyl carnitines, andsalts and mixtures thereof.

Hydrophilic non-ionic surfactants may include, but not limited to,alkylglucosides; alkylmaltosides; alkylthioglucosides; laurylmacrogolglycerides; polyoxyalkylene alkyl ethers such as polyethyleneglycol alkyl ethers; polyoxyalkylene alkylphenols such as polyethyleneglycol alkyl phenols; polyoxyalkylene alkyl phenol fatty acid esterssuch as polyethylene glycol fatty acids monoesters and polyethyleneglycol fatty acids diesters; polyethylene glycol glycerol fatty acidesters; polyglycerol fatty acid esters; polyoxyalkylene sorbitan fattyacid esters such as polyethylene glycol sorbitan fatty acid esters;hydrophilic transesterification products of a polyol with at least onemember of the group consisting of glycerides, vegetable oils,hydrogenated vegetable oils, fatty acids, and sterols; polyoxyethylenesterols, derivatives, and analogues thereof; polyoxyethylated vitaminsand derivatives thereof; polyoxyethylene-polyoxypropylene blockcopolymers; and mixtures thereof; polyethylene glycol sorbitan fattyacid esters and hydrophilic transesterification products of a polyolwith at least one member of the group consisting of triglycerides,vegetable oils, and hydrogenated vegetable oils. The polyol may beglycerol, ethylene glycol, polyethylene glycol, sorbitol, propyleneglycol, pentaerythritol, or a saccharide.

Other hydrophilic-non-ionic surfactants include, without limitation,PEG-10 laurate, PEG-12 laurate, PEG-20 laurate, PEG-32 laurate, PEG-32dilaurate, PEG-12 oleate, PEG-15 oleate, PEG-20 oleate, PEG-20 dioleate,PEG-32 oleate, PEG-200 oleate, PEG-400 oleate, PEG-15 stearate, PEG-32distearate, PEG-40 stearate, PEG-100 stearate, PEG-20 dilaurate, PEG-25glyceryl trioleate, PEG-32 dioleate, PEG-20 glyceryl laurate, PEG-30glyceryl laurate, PEG-20 glyceryl stearate, PEG-20 glyceryl oleate,PEG-30 glyceryl oleate, PEG-30 glyceryl laurate, PEG-40 glyceryllaurate, PEG-40 palm kernel oil, PEG-50 hydrogenated castor oil, PEG-40castor oil, PEG-35 castor oil, PEG-60 castor oil, PEG-40 hydrogenatedcastor oil, PEG-60 hydrogenated castor oil, PEG-60 corn oil, PEG-6caprate/caprylate glycerides, PEG-8 caprate/caprylate glycerides,polyglyceryl-10 laurate, PEG-30 cholesterol, PEG-25 phyto sterol, PEG-30soya sterol, PEG-20 trioleate, PEG-40 sorbitan oleate, PEG-80 sorbitanlaurate, polysorbate 20, polysorbate 80, POE-9 lauryl ether, POE-23lauryl ether, POE-10 oleyl ether, POE-20 oleyl ether, POE-20 stearylether, tocopheryl PEG-100 succinate, PEG-24 cholesterol,polyglyceryl-10oleate, Tween 40, Tween 60, sucrose monostearate, sucrosemonolaurate, sucrose monopalmitate, PEG 10-100 nonyl phenol series, PEG15-100 octyl phenol series, and poloxamers.

Suitable lipophilic surfactants include, by way of example only: fattyalcohols, glycerol fatty acid esters, acetylated glycerol fatty acidesters, lower alcohol fatty acids esters, propylene glycol fatty acidesters, sorbitan fatty acid esters, polyethylene glycol sorbitan fattyacid esters, sterols and sterol derivatives, polyoxyethylated sterolsand sterol derivatives, polyethylene glycol alkyl ethers, sugar esters,sugar ethers, lactic acid derivatives of mono- and di-glycerides, andhydrophobic transesterification products of a polyol with at least onemember of the group consisting of glycerides, vegetable oils,hydrogenated vegetable oils, fatty acids and sterols, oil-solublevitamins/vitamin derivatives, and mixtures thereof. Within this group,preferred lipophilic surfactants include glycerol fatty acid esters,propylene glycol fatty acid esters, and mixtures thereof, or arehydrophobic transesterification products of a polyol with at least onemember of the group consisting of vegetable oils, hydrogenated vegetableoils, and triglycerides.

In an embodiment, the composition may include a solubilizer to ensuregood solubilization and/or dissolution of the compound of the presentinvention and to minimize precipitation of the compound of the presentinvention. This can be especially important for compositions fornon-oral use—e.g., compositions for injection. A solubilizer may also beadded to increase the solubility of the hydrophilic drug and/or othercomponents, such as surfactants, or to maintain the composition as astable or homogeneous solution or dispersion.

Examples of suitable solubilizers include, but are not limited to, thefollowing: alcohols and polyols, such as ethanol, isopropanol, butanol,benzyl alcohol, ethylene glycol, propylene glycol, butanediols andisomers thereof, glycerol, pentaerythritol, sorbitol, mannitol, xylitol,transcutol, dimethyl isosorbide, polyethylene glycol, polypropyleneglycol, polyvinylalcohol, hydroxypropyl methylcellulose and othercellulose derivatives, cyclodextrins and cyclodextrin derivatives;ethers of polyethylene glycols having an average molecular weight ofabout 200 to about 6000, such as tetrahydrofurfuryl alcohol PEG ether(glycofurol) or methoxy PEG; amides and other nitrogen-containingcompounds such as 2-pyrrolidone, 2-piperidone, ε-caprolactam,N-alkylpyrrolidone, N-hydroxyalkylpyrrolidone, N-alkylpiperidone,N-alkylcaprolactam, dimethylacetamide and polyvinylpyrrolidone; esterssuch as ethyl propionate, tributylcitrate, acetyl triethylcitrate,acetyl tributyl citrate, triethylcitrate, ethyl oleate, ethyl caprylate,ethyl butyrate, triacetin, propylene glycol monoacetate, propyleneglycol diacetate, epsilon-caprolactone and isomers thereof,δ-valerolactone and isomers thereof, β-butyrolactone and isomersthereof; and other solubilizers known in the art, such as dimethylacetamide, dimethyl isosorbide, N-methyl pyrrolidones, monooctanoin,diethylene glycol monoethyl ether, and water.

Mixtures of solubilizers may also be used. Examples include, but notlimited to, triacetin, triethylcitrate, ethyl oleate, ethyl caprylate,dimethylacetamide, N-methylpyrrolidone, N-hydroxyethylpyrrolidone,polyvinylpyrrolidone, hydroxypropyl methylcellulose, hydroxypropylcyclodextrins, ethanol, polyethylene glycol 200-100, glycofurol,transcutol, propylene glycol, and dimethyl isosorbide. Particularlypreferred solubilizers include sorbitol, glycerol, triacetin, ethylalcohol, PEG-400, glycofurol and propylene glycol.

The amount of solubilizer that can be included is not particularlylimited. The amount of a given solubilizer may be limited to abioacceptable amount, which may be readily determined by one of skill inthe art. In some circumstances, it may be advantageous to includeamounts of solubilizers far in excess of bioacceptable amounts, forexample to maximize the concentration of the drug, with excesssolubilizer removed prior to providing the composition to a patientusing conventional techniques, such as distillation or evaporation.Thus, if present, the solubilizer can be in a weight ratio of 10%, 25%,50%, 100%, or up to about 200% by weight, based on the combined weightof the drug, and other excipients. If desired, very small amounts ofsolubilizer may also be used, such as 5%, 2%, 1% or even less.Typically, the solubilizer may be present in an amount of about 1% toabout 100%, more typically about 5% to about 25% by weight.

The composition can further include one or more pharmaceuticallyacceptable additives and excipients. Such additives and excipientsinclude, without limitation, detackifiers, anti-foaming agents,buffering agents, polymers, antioxidants, preservatives, chelatingagents, viscomodulators, tonicifiers, flavorants, colorants, odorants,opacifiers, suspending agents, binders, fillers, plasticizers,lubricants, and mixtures thereof.

In addition, an acid or a base may be incorporated into thepharmaceutical composition to facilitate processing, to enhancestability, or for other reasons. Examples of pharmaceutically acceptablebases include amino acids, amino acid esters, ammonium hydroxide,potassium hydroxide, sodium hydroxide, sodium hydrogen carbonate,aluminum hydroxide, calcium carbonate, magnesium hydroxide, magnesiumaluminum silicate, synthetic aluminum silicate, synthetic hydrocalcite,magnesium aluminum hydroxide, diisopropylethylamine, ethanolamine,ethylenediamine, triethanolamine, triethylamine, triisopropanolamine,trimethylamine, tris(hydroxymethyl)aminomethane (TRIS) and the like.Also suitable are bases that are salts of a pharmaceutically acceptableacid, such as acetic acid, acrylic acid, adipic acid, alginic acid,alkanesulfonic acid, amino acids, ascorbic acid, benzoic acid, boricacid, butyric acid, carbonic acid, citric acid, fatty acids, formicacid, fumaric acid, gluconic acid, hydroquinosulfonic acid, isoascorbicacid, lactic acid, maleic acid, oxalic acid, para-bromophenylsulfonicacid, propionic acid, p-toluenesulfonic acid, salicylic acid, stearicacid, succinic acid, tannic acid, tartaric acid, thioglycolic acid,toluenesulfonic acid, uric acid, and the like. Salts of polyproticacids, such as sodium phosphate, disodium hydrogen phosphate, and sodiumdihydrogen phosphate can also be used. When the base is a salt, thecation can be any convenient and pharmaceutically acceptable cation,such as ammonium, alkali metals and alkaline earth metals. Example mayinclude, but not limited to, sodium, potassium, lithium, magnesium,calcium and ammonium.

Suitable acids are pharmaceutically acceptable organic or inorganicacids. Examples of suitable inorganic acids include hydrochloric acid,hydrobromic acid, hydriodic acid, sulfuric acid, nitric acid, boricacid, phosphoric acid, and the like. Examples of suitable organic acidsinclude acetic acid, acrylic acid, adipic acid, alginic acid,alkanesulfonic acids, amino acids, ascorbic acid, benzoic acid, boricacid, butyric acid, carbonic acid, citric acid, fatty acids, formicacid, fumaric acid, gluconic acid, hydroquinosulfonic acid, isoascorbicacid, lactic acid, maleic acid, methanesulfonic acid, oxalic acid,para-bromophenylsulfonic acid, propionic acid, p-toluenesulfonic acid,salicylic acid, stearic acid, succinic acid, tannic acid, tartaric acid,thioglycolic acid, toluenesulfonic acid and uric acid.

Dosages and Dosing Regimens

In an embodiment, the invention provides a pharmaceutical compositioncomprising a crystalline form of the free base of Formula (1). In anembodiment, the invention provides a pharmaceutical compositioncomprising a crystalline solvate of the free base of Formula (1). In anembodiment, the invention provides a pharmaceutical compositioncomprising a crystalline hydrate of the free base of Formula (1). In anembodiment, the invention provides a pharmaceutical compositioncomprising a crystalline salt of Formula (1). In an embodiment, theinvention provides a pharmaceutical composition comprising an amorphousform of Formula (1).

The amounts of the solid form of the BTK inhibitor of Formula (1), willbe dependent on the mammal being treated, the severity of the disorderor condition, the rate of administration, the disposition of thecompounds and the discretion of the prescribing physician. However, aneffective dosage is in the range of about 0.001 to about 100 mg per kgbody weight per day, such as about 1 to about 35 mg/kg/day, in single ordivided doses. For a 70 kg human, this would amount to about 0.05 to 7g/day, such as about 0.05 to about 2.5 g/day. In some instances, dosagelevels below the lower limit of the aforesaid range may be more thanadequate, while in other cases still larger doses may be employedwithout causing any harmful side effect, for example by dividing suchlarger doses into several small doses for administration throughout theday.

In selected embodiments, a solid form of the BTK inhibitor of Formula(1) is administered in a single dose. Typically, such administrationwill be by injection, for example by intravenous injection, in order tointroduce the active pharmaceutical ingredients quickly. However, otherroutes may be used as appropriate. A single dose of a solid form of theBTK inhibitor of Formula (1) may also be used for treatment of an acutecondition.

In selected embodiments, a solid form of the BTK inhibitor of Formula(1) is administered in multiple doses. Dosing may be about once, twice,three times, four times, five times, six times, or more than six timesper day. Dosing may be about once a month, once every two weeks, once aweek, or once every other day. In other embodiments, a solid form of theBTK inhibitor of Formula (1) is administered about once per day to about6 times per day. In another embodiment the administration of the BTKinhibitor of Formula (1), continues for less than about 7 days. In yetanother embodiment the administration continues for more than about 6,10, 14, 28 days, two months, six months, or one year. In some cases,continuous dosing is achieved and maintained as long as necessary.

Administration of the active pharmaceutical ingredients of the inventionmay continue as long as necessary. In selected embodiments, a solid formof the BTK inhibitor of Formula (1), are administered for more than 1,2, 3, 4, 5, 6, 7, 14, or 28 days. In some embodiments, the solid formsof the BTK inhibitor of Formula (1) are administered for less than 28,14, 7, 6, 5, 4, 3, 2, or 1 day. In selected embodiments, a solid form ofthe BTK inhibitor of Formula (1) is administered chronically on anongoing basis—e.g., for the treatment of chronic effects.

In some embodiments, an effective dosage of a solid form of the BTKinhibitor of Formula (1) is in the range of about 1 mg to about 500 mg,about 10 mg to about 300 mg, about 20 mg to about 250 mg, about 25 mg toabout 200 mg, about 10 mg to about 200 mg, about 20 mg to about 150 mg,about 30 mg to about 120 mg, about 10 mg to about 90 mg, about 20 mg toabout 80 mg, about 30 mg to about 70 mg, about 40 mg to about 60 mg,about 45 mg to about 55 mg, about 48 mg to about 52 mg, about 50 mg toabout 150 mg, about 60 mg to about 140 mg, about 70 mg to about 130 mg,about 80 mg to about 120 mg, about 90 mg to about 110 mg, about 95 mg toabout 105 mg, about 150 mg to about 250 mg, about 160 mg to about 240mg, about 170 mg to about 230 mg, about 180 mg to about 220 mg, about190 mg to about 210 mg, about 195 mg to about 205 mg, or about 198 toabout 202 mg. In some embodiments, an effective dosage of a solid formof the BTK inhibitor of Formula (1) is about 25 mg, about 50 mg, about75 mg, about 100 mg, about 125 mg, about 150 mg, about 175 mg, about 200mg, about 225 mg, about 250 mg, about 275 mg, about 300 mg, about 325mg, about 350 mg, about 375 mg, about 400 mg, about 425 mg, about 450mg, about 475 mg, or about 500 mg. In some embodiments, an effectivedosage of a solid form of the BTK inhibitor of Formula (1) is 25 mg, 50mg, 75 mg, 100 mg, 125 mg, 150 mg, 175 mg, 200 mg, 225 mg, 250 mg, 275mg, 300 mg, 325 mg, 350 mg, 375 mg, 400 mg, 425 mg, 450 mg, 475 mg, or500 mg.

In some embodiments, an effective dosage of a solid form of the BTKinhibitor of Formula (1) is in the range of about 0.01 mg/kg to about4.3 mg/kg, about 0.15 mg/kg to about 3.6 mg/kg, about 0.3 mg/kg to about3.2 mg/kg, about 0.35 mg/kg to about 2.85 mg/kg, about 0.15 mg/kg toabout 2.85 mg/kg, about 0.3 mg to about 2.15 mg/kg, about 0.45 mg/kg toabout 1.7 mg/kg, about 0.15 mg/kg to about 1.3 mg/kg, about 0.3 mg/kg toabout 1.15 mg/kg, about 0.45 mg/kg to about 1 mg/kg, about 0.55 mg/kg toabout 0.85 mg/kg, about 0.65 mg/kg to about 0.8 mg/kg, about 0.7 mg/kgto about 0.75 mg/kg, about 0.7 mg/kg to about 2.15 mg/kg, about 0.85mg/kg to about 2 mg/kg, about 1 mg/kg to about 1.85 mg/kg, about 1.15mg/kg to about 1.7 mg/kg, about 1.3 mg/kg mg to about 1.6 mg/kg, about1.35 mg/kg to about 1.5 mg/kg, about 2.15 mg/kg to about 3.6 mg/kg,about 2.3 mg/kg to about 3.4 mg/kg, about 2.4 mg/kg to about 3.3 mg/kg,about 2.6 mg/kg to about 3.15 mg/kg, about 2.7 mg/kg to about 3 mg/kg,about 2.8 mg/kg to about 3 mg/kg, or about 2.85 mg/kg to about 2.95mg/kg. In some embodiments, an effective dosage of a solid form of theBTK inhibitor of Formula (1) is about 0.35 mg/kg, about 0.7 mg/kg, about1 mg/kg, about 1.4 mg/kg, about 1.8 mg/kg, about 2.1 mg/kg, about 2.5mg/kg, about 2.85 mg/kg, about 3.2 mg/kg, or about 3.6 mg/kg.

In some embodiments, a solid form of the BTK inhibitor of Formula (1) isadminstered at a dosage of 10 to 400 mg once daily (QD), including adosage of 5 mg, 10 mg, 12.5 mg, 25 mg, 50 mg, 75 mg, 100 mg, 150 mg, 175mg, 200 mg, 225 mg, 250 mg, 275 mg, 300 mg, 325 mg, 350 mg, 375 mg, 400mg, 425 mg, 450 mg, 475 mg, and 500 mg once daily (QD).

In some embodiments, a solid form of the BTK inhibitor of Formula (1) isadminstered at a dosage of 10 to 400 mg BID, including a dosage of 5 mg,10 mg, 12.5 mg, 25 mg, 50 mg, 75 mg, 100 mg, 150 mg, 175 mg, 200 mg, 225mg, 250 mg, 275 mg, 300 mg, 325 mg, 350 mg, 375 mg, 400 mg, 425 mg, 450mg, 475 mg, and 500 mg BID.

In some embodiments, a solid form of the BTK inhibitor of Formula (1) isadministered at a dosage of 10 to 400 mg TID, including a dosage of 5mg, 10 mg, 12.5 mg, 25 mg, 50 mg, 75 mg, 100 mg, 150 mg, 175 mg, 200 mg,225 mg, 250 mg, 275 mg, 300 mg, 325 mg, 350 mg, 375 mg, 400 mg, 425 mg,450 mg, 475 mg, and 500 mg TID.

An effective amount of a solid form of the BTK inhibitors may beadministered in either single or multiple doses by any of the acceptedmodes of administration of active pharmaceutical ingredients havingsimilar utilities, including rectal, buccal, intranasal and transdermalroutes, by intra-arterial injection, intravenously, intraperitoneally,parenterally, intramuscularly, subcutaneously, orally, topically, or asan inhalant.

Pharmaceutical Compositions for Overcoming the Effects of Acid ReducingAgents

The compositions and methods described herein can be used to overcomethe effects of acid reducing agents. Acid-reducing agents can greatlylimit the exposure of weakly acidic drugs (such as Formula (1) freebase) in mammals. Smelick, et al., Mol. Pharmaceutics 2013, 10,4055-4062. Acid reducing agents include proton pump inhibitors, such asomeprazole, esomeprazole, lansoprazole, dexlansoprazole, pantoprazole,rabeprazole, and ilaprazole; H₂ receptor antagonists, such ascimetidine, ranitidine, and famotidine; and antacids such asbicarbonates, carbonates, and hydroxides of aluminium, calcium,magnesium, potassium, and sodium, as well as mixtures of antacids withagents targeting mechanisms of gastric secretion. Overcoming the effectsof acid reducing agents is a significant issue in the treatment ofpatients with cancer, inflammatory diseases, immune diseases, andautoimmune diseases, since these patients are commonly co-administeredacid reducing agents for gastric irritation that often accompanies theirconditions. Acid reducing agents are the most commonly prescribedmedications in North America and Western Europe. Most recently approvedoral cancer therapeutics have pH-dependent solubility and thus apotential drug-drug interaction with regards to acid reducing agents. Incancer patients, it is estimated that 20-33% of all patients are usingsome form of acid-reducing agent. In particular cancers, such aspancreatic cancer or gastrointestinal cancers, acid reducing agent useis as high as 60-80% of patients. Smelick, et al., Mol. Pharmaceutics2013, 10, 4055-4062.

In an embodiment, a pharmaceutical composition comprises a BTK inhibitoraccording to Formula (1) and an acidulant. In an embodiment, apharmaceutical composition comprises a BTK inhibitor according toFormula (1) and an acidulant selected from the group consisting offumaric acid, tartaric acid, ascorbic acid, alginic acid, sodiumalginate, potassium alginate, and Carbopol 971P (carboxypolymethylene).In an embodiment, a pharmaceutical composition comprises a BTK inhibitoraccording to Formula (1) and an acidulant selected from the groupconsisting of fumaric acid, succinic acid, D-tartaric acid, L-tartaricacid, racemic tartaric acid, ascorbic acid, isoascorbic acid (also knownas erythorbic acid and D-araboascorbic acid), alginic acid, Protacid F120 NM, Protacid AR 1112 (also known as Kelacid NF), Carbomer 941(polyacrylic acid), and Carbopol 971P (carboxypolymethylene). In anembodiment, the solid form of Formula (1) in any of the foregoingembodiments is Form I of the free base. In an embodiment, the acidulantis extragranular. In an embodiment, the acidulant is intragranular.

Alginic acid is a polysaccharide copolymer, β-D-mannuronic acid (M) andα-L-guluronic acid (G) linked by 1-4 glycosidic bonds. In an embodiment,a pharmaceutical composition comprises a BTK inhibitor according toFormula (1) and an acidulant that is an alginic acid, or a salt thereof,wherein the alginic acid, or a salt thereof, exhibits an M/G ratioselected from the group consisting of between 0.1 and 0.5, between 0.2and 0.6, between 0.3 and 0.7, between 0.4 and 0.8, between 0.5 and 0.9,between 0.6 and 1.0, between 0.7 and 1.1, between 0.8 and 1.2, between0.9 and 1.3, between 1.0 and 1.4, between 1.1 and 1.5, between 1.2 and1.6, between 1.3 and 1.7, between 1.4 and 1.8, between 1.5 and 1.9,between 1.6 and 2.0, between 1.7 and 2.1, between 1.8 and 2.2, between1.9 and 2.3, between 2.0 and 2.4, and between 2.1 and 2.5. In anembodiment, a pharmaceutical composition comprises a BTK inhibitoraccording to Formula (1) and an acidulant that is an alginic acid, or asalt thereof, wherein the alginic acid, or a salt thereof, exhibits anM/G ratio selected from the group consisting of less than 0.5, less than1.0, less than 1.5, less than 2.0, and less than 2.5. In an embodiment,a pharmaceutical composition comprises a BTK inhibitor according toFormula (1) and an acidulant that is an alginic acid, or a salt thereof,wherein the alginic acid, or a salt thereof, exhibits an M/G ratioselected from the group consisting of greater than 0.5, greater than1.0, greater than 1.5, greater than 2.0, and greater than 2.5. In anembodiment, a pharmaceutical composition comprises a BTK inhibitoraccording to Formula (1) and an acidulant that is an alginic acid, or asalt thereof, wherein the alginic acid, or a salt thereof, exhibits anM/G ratio selected from the group consisting of 0.1, 0.2, 0.3, 0.4, 0.5,0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9,2.0, 2.1, 2.2, 2.3, 2.4, and 2.5. In an embodiment, the solid form ofFormula (1) in any of the foregoing embodiments is Form I of the freebase. M/G ratio, as well as the fraction of M and G groups, thefractions of MM and GG “diads,” the fractions of “triads” (e.g., MGG),and the fractions of larger sequences of M and G groups, may bedetermined by methods known to those of ordinary skill in the art,including nuclear magnetic resonance (NMR) spectroscopy (with ourwithout digestion) and mass spectrometry. Larsen, et al., Carbohydr.Res., 2003, 338, 2325-2336.

In an embodiment, a pharmaceutical composition comprises a BTK inhibitoraccording to Formula (1) and an acidulant in a concentration (% mass)selected from the group consisting of between 1% and 5%, between 5% and10%, between 10% and 15%, between 15% and 20%, between 20% and 25%,between 25% and 30%, and between 30% and 35%. In an embodiment, apharmaceutical composition comprises a BTK inhibitor according toFormula (1) and an acidulant in a concentration (% mass) selected fromthe group consisting of between 1% and 5%, between 5% and 10%, between10% and 15%, between 15% and 20%, between 20% and 25%, between 25% and30%, and between 30% and 35%, wherein the acidulant is selected from thegroup consisting of fumaric acid, succinic acid, D-tartaric acid,L-tartaric acid, racemic tartaric acid, ascorbic acid, isoascorbic acid(also known as erythorbic acid and D-araboascorbic acid), alginic acid,sodium alginate, potassium alginate, Protacid F 120 NM, Protacid AR 1112(also known as Kelacid NF), and Carbopol 971P (carboxypolymethylene). Inan embodiment, the solid form of Formula (1) in any of the foregoingembodiments is Form I of the free base.

In an embodiment, a pharmaceutical composition comprises a BTK inhibitoraccording to Formula (1) and an acidulant in a concentration (% mass)selected from the group consisting of less than 1%, less than 5%, lessthan 10%, less than 15%, less than 20%, less than 25%, less than 30%,and less than 35%. In an embodiment, a pharmaceutical compositioncomprises a BTK inhibitor according to Formula (1) and an acidulant in aconcentration (% mass) selected from the group consisting of less than1%, less than 5%, less than 10%, less than 15%, less than 20%, less than25%, less than 30%, and less than 35%, wherein the acidulant is selectedfrom the group consisting of fumaric acid, succinic acid, D-tartaricacid, L-tartaric acid, racemic tartaric acid, ascorbic acid, isoascorbicacid (also known as erythorbic acid and D-araboascorbic acid), alginicacid, sodium alginate, potassium alginate, Protacid F 120 NM, ProtacidAR 1112 (also known as Kelacid NF), and Carbopol 971P(carboxypolymethylene). In an embodiment, the solid form of Formula (1)in any of the foregoing embodiments is Form I of the free base.

In an embodiment, a pharmaceutical composition comprises a BTK inhibitoraccording to Formula (1) and an acidulant in a concentration (% mass)selected from the group consisting of greater than 1%, greater than 5%,greater than 10%, greater than 15%, greater than 20%, greater than 25%,greater than 30%, and greater than 35%. In an embodiment, apharmaceutical composition comprises a BTK inhibitor according toFormula (1) and an acidulant in a concentration (% mass) selected fromthe group consisting of greater than 1%, greater than 5%, greater than10%, greater than 15%, greater than 20%, greater than 25%, greater than30%, and greater than 35%, wherein the acidulant is selected from thegroup consisting of fumaric acid, succinic acid, D-tartaric acid,L-tartaric acid, racemic tartaric acid, ascorbic acid, isoascorbic acid(also known as erythorbic acid and D-araboascorbic acid), alginic acid,sodium alginate, potassium alginate, Protacid F 120 NM, Protacid AR 1112(also known as Kelacid NF), and Carbopol 971P (carboxypolymethylene). Inan embodiment, the solid form of Formula (1) in any of the foregoingembodiments is Form I of the free base.

In an embodiment, a pharmaceutical composition comprises a BTK inhibitoraccording to Formula (1) and an acidulant in a concentration (% mass)selected from the group consisting of about 1%, about 2%, about 3%,about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%,about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%,about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%,about 37%, about 38%, about 39%, and about 40%. In an embodiment, apharmaceutical composition comprises a BTK inhibitor according toFormula (1) and an acidulant in a concentration (% mass) selected fromthe group consisting of about 1%, about 2%, about 3%, about 4%, about5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%,about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%,about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about38%, about 39%, and about 40%, wherein the acidulant is selected fromthe group consisting of fumaric acid, succinic acid, D-tartaric acid,L-tartaric acid, racemic tartaric acid, ascorbic acid, isoascorbic acid(also known as erythorbic acid and D-araboascorbic acid), alginic acid,sodium alginate, potassium alginate, Protacid F 120 NM, Protacid AR 1112(also known as Kelacid NF), and Carbopol 971P (carboxypolymethylene). Inan embodiment, the solid form of Formula (1) in any of the foregoingembodiments is Form I of the free base.

In an embodiment, a pharmaceutical composition comprises a BTK inhibitoraccording to Formula (1) and an extragranular acidulant, wherein theextragranular acidulant is selected from the group consisting of fumaricacid, succinic acid, D-tartaric acid, L-tartaric acid, racemic tartaricacid, ascorbic acid, isoascorbic acid (also known as erythorbic acid andD-araboascorbic acid), alginic acid, sodium alginate, potassiumalginate, Protacid F 120 NM, Protacid AR 1112 (also known as KelacidNF), and Carbopol 971P (carboxypolymethylene), and combinations thereof.In an embodiment, a pharmaceutical composition comprises a BTK inhibitoraccording to Formula (1) and an extragranular acidulant, wherein theextragranular acidulant is fumaric acid at a concentration of betweenabout 15% to about 33% by weight. In an embodiment, a pharmaceuticalcomposition comprises a BTK inhibitor according to Formula (1) and anextragranular acidulant, wherein the extragranular acidulant is alginicacid or a salt thereof (such as sodium alginate or potassium alginate)at a concentration of between about 5% to about 33% by weight. In anembodiment, a pharmaceutical composition comprises a BTK inhibitoraccording to Formula (1) and an extragranular acidulant, wherein theextragranular acidulant is L-tartaric acid at a concentration of betweenabout 25% to about 33% by weight. In an embodiment, a pharmaceuticalcomposition comprises a BTK inhibitor according to Formula (1) and anextragranular acidulant, wherein the extragranular acidulant is ascorbicacid at a concentration of between about 20% to about 50% by weight andCarbopol 971P (carboxypolymethylene) at a concentration of between about2.5% to about 10% by weight. In an embodiment, a pharmaceuticalcomposition comprises a BTK inhibitor according to Formula (1) and anextragranular acidulant, wherein the extragranular acidulant is fumaricacid at a concentration of between about 5% to about 15% by weight andalginic acid or a salt thereof at a concentration of about 15% to about33% by weight. In an embodiment, a pharmaceutical composition comprisesa BTK inhibitor according to Formula (1) and an extragranular acidulant,wherein the extragranular acidulant is L-tartaric acid at aconcentration of between about 5% to 15% by weight and alginic acid at aconcentration of between about 15% to about 33% by weight.

In an embodiment, a pharmaceutical composition comprises a BTK inhibitoraccording to Formula (1) and an acidulant, wherein the acidulent isselected from the group consisting of fumaric acid, maleic acid,phosphoric acid, L-tartaric acid, citric acid, gentisic acid, oxalicacid, and sulfuric acid. In an embodiment, a pharmaceutical compositioncomprises a BTK inhibitor according to Formula (1) and an acidulant,wherein the acidulent is selected from the group consisting of fumaricacid, maleic acid, phosphoric acid, L-tartaric acid, citric acid,gentisic acid, oxalic acid, and sulfuric acid, and wherein the acidulantis a salt counterion included in a single crystalline phase with Formula(1).

In an embodiment, in addition to an acidulant, a pharmaceuticalcomposition includes an excipient to prolong the exposure of Formula (1)to the acidic microenvironment. In an embodiment, this excipient is apolymer of natural, synthetic or semisynthetic origins. The polymer maycontain acidic, anionic, or non-ionic monomers, oligomers or polymers ora mixture of acidic, anionic and non-ionic monomers or copolymers. Inone version the excipient is selected from the group consisting ofhydroxypropylmethylcellulose, low substituted hydroxypropylcellulose,hydroxypropylcellulose, tocopherol polyethyleneoxide succinate(D-α-tocopherol polyethylene glycol succinate, TPGS, or vitamin E TPGS),methylcellulose, comboxymethylcellulose, sodium carboxymethylcellulose,methylacrylate, ethylacrylate, co-polymers of methyl and ethyl acrylate,hydroxypropylmethylcellulose acetate succinate, gelatin, maize starch,pea starch, modified maize starch, potato starch, modified potatostarch, sodium starch glycolate, croscarmellose, crospovidone,copovidone, polyethylene glycol, polypropylene glycol, polyethylene andpolypropylene glycol copolymers, polyvinylalcohol, polyvinylalcohol andpolyethylene oxide copolymers. Copolymers of the foregoing polymers,where applicable, may also be used. Copolymers may be block, branched orterminal copolymers. In an embodiment, the polymer exhibits swelling,binding, or gelling properties that inhibit the disintegration,dissolution, and erosion of the pharmaceutical composition in order toprolong dissolution or to increase total dissolution. In an embodiment,the inclusion of the polymer increases dissolution rate and extent ofdissolution over the use of an acidulant alone. The swelling, binding orgelling properties are pH-dependant in one embodiment, wherein thepolymer swells, binds, or gels at one pH or range of pH in a differentmanner than at another pH. In one embodiment this may decreasedissolution at a lower pH than at a higher pH or vice versa. In anotherembodiment this leads to similar dissolution of Formula I in acidic,neutral or basic pH. This leads to similar plasma exposure independentof stomach pH.

The dissolution profile of a formulation containing one or moreswelling, gelling, or binding excipients may exhibit a zero, first, orsecond differential rate order at one or more pH value or a mixture ofdifferent rate orders at different pH values. In an embodiment, apharmaceutical composition will provide a constant level of drug intothe gastrointestinal tract of a mammal by dissolution. Where the Formula(1) is absorbed, this leads to a sustained plasma level of drug over aperiod, delays the t_(max), and reduces the c_(max) of an equivalentdose of an immediate release formulation of Formula (1). In anotherembodiment this leads to similar exposure in a mammal regardless ofstomach pH.

Methods of Treating Solid Tumor Cancers, Hematological Malignancies,Inflammatory Diseases, Autoimmune Disorders, Immune Disorders, and OtherDiseases

The pharmaceutical compositions described herein can be used in a methodfor treating diseases. In preferred embodiments, they are for use intreating hyperproliferative disorders. They may also be used in treatingother disorders as described herein and in the following paragraphs.

In some embodiments, the invention provides a method of treating ahyperproliferative disorder in a mammal that comprises administering tothe mammal a therapeutically effective amount of a crystalline oramorphous solid form of Formula (1), or a pharmaceutical compositioncomprising a crystalline or amorphous solid form of Formula (1), asdescribed herein each including Form I of the free base of Formula (1).In some embodiments, the hyperproliferative disorder is cancer. Inpreferred embodiments, the cancer is selected from the group consistingof chronic lymphocytic leukemia, non-Hodgkin's lymphoma, diffuse largeB-cell lymphoma, mantle cell lymphoma, follicular lymphoma, andWaldenström's macroglobulinemia. In preferred embodiments, the cancer isselected from the group consisting of non-Hodgkin's lymphomas (such asdiffuse large B-cell lymphoma), acute myeloid leukemia, thymus, brain,lung, squamous cell, skin, eye, retinoblastoma, intraocular melanoma,oral cavity and oropharyngeal, bladder, gastric, stomach, pancreatic,bladder, breast, cervical, head, neck, renal, kidney, liver, ovarian,prostate, colorectal, bone (e.g., metastatic bone), esophageal,testicular, gynecological, thyroid, CNS, PNS, AIDS-related (e.g.,lymphoma and Kaposi's sarcoma), viral-induced cancers such as cervicalcarcinoma (human papillomavirus), B-cell lymphoproliferative disease andnasopharyngeal carcinoma (Epstein-Barr virus), Kaposi's sarcoma andprimary effusion lymphomas (Kaposi's sarcoma herpesvirus),hepatocellular carcinoma (hepatitis B and hepatitis C viruses), andT-cell leukemias (Human T-cell leukemia virus-1), B cell acutelymphoblastic leukemia, Burkitt's leukemia, juvenile myelomonocyticleukemia, hairy cell leukemia, Hodgkin's disease, multiple myeloma, mastcell leukemia, and mastocytosis. In selected embodiments, the methodrelates to the treatment of a non-cancerous hyperproliferative disordersuch as benign hyperplasia of the skin (e.g., psoriasis), restenosis, orprostate conditions (e.g., benign prostatic hypertrophy (BPH)). In someembodiments, the hyperproliferative disorder is an inflammatory, immune,or autoimmune disorder. In some embodiments, the hyperproliferativedisorder is selected from the group consisting of tumor angiogenesis,chronic inflammatory disease, rheumatoid arthritis, atherosclerosis,inflammatory bowel disease, skin diseases such as psoriasis, eczema, andscleroderma, diabetes, diabetic retinopathy, retinopathy of prematurity,age-related macular degeneration, hemangioma, glioma and melanoma,ulcerative colitis, atopic dermatitis, pouchitis, spondylarthritis,uveitis, Behcet's disease, polymyalgia rheumatica, giant-cell arteritis,sarcoidosis, Kawasaki disease, juvenile idiopathic arthritis,hidratenitis suppurativa, Sjögren's syndrome, psoriatic arthritis,juvenile rheumatoid arthritis, ankylosing spondylitis, Crohn's disease,lupus, and lupus nephritis. In an embodiment, the solid form of Formula(1) in any of the foregoing embodiments is Form I of the free base. Inan embodiment, the method of any of the foregoing embodiments furtherincludes the step of administering an acid reducing agent to the mammal.In an embodiment, the acid reducing agent is selected from the groupconsisting of proton pump inhibitors, such as omeprazole, esomeprazole,lansoprazole, dexlansoprazole, pantoprazole, rabeprazole, andilaprazole; H₂ receptor antagonists, such as cimetidine, ranitidine, andfamotidine; and antacids such as bicarbonates, carbonates, andhydroxides of aluminium, calcium, magnesium, potassium, and sodium.

In some embodiments, the invention provides pharmaceutical compositionsof a solid form of Formula (1) described herein for use in the treatmentof cancers such as thymus cancer, brain cancer (e.g., glioma), lungcancer, squamous cell cancer, skin cancer (e.g., melanoma), eye cancer,retinoblastoma cancer, intraocular melanoma cancer, oral cavity cancer,oropharyngeal cancer, bladder cancer, gastric cancer, stomach cancer,pancreatic cancer, bladder cancer, breast cancer, cervical cancer, headand neck cancer, renal cancer, kidney cancer, liver cancer, ovariancancer, prostate cancer, colorectal cancer, colon cancer, esophagealcancer, testicular cancer, gynecological cancer, ovarian cancer, thyroidcancer, CNS cancer, PNS cancer, AIDS-related cancer (e.g., lymphoma andKaposi's sarcoma), viral-induced cancer, and epidermoid cancer. In someembodiments, the invention provides pharmaceutical compositions of asolid form of Formula (1) described herein for the treatment of anon-cancerous hyperproliferative disorder such as benign hyperplasia ofthe skin (e.g., psoriasis), restenosis, or prostate (e.g., benignprostatic hypertrophy (BPH)). In some embodiments, the inventionprovides pharmaceutical compositions of a solid form of Formula (1)described herein for use in the treatment of disorders such asmyeloproliferative disorders (MPDs), myeloproliferative neoplasms,polycythemia vera (PV), essential thrombocythemia (ET), primarymyelofibrosis (PMF), myelodysplastic syndrome, chronic myelogenousleukemia (BCR-ABL1-positive), chronic neutrophilic leukemia, chroniceosinophilic leukemia, or mastocytosis. The invention also providescompositions for use in treating a disease related to vasculogenesis orangiogenesis in a mammal which can manifest as tumor angiogenesis,chronic inflammatory disease such as rheumatoid arthritis, inflammatorybowel disease, atherosclerosis, skin diseases such as psoriasis, eczema,and scleroderma, diabetes, diabetic retinopathy, retinopathy ofprematurity, age-related macular degeneration, and hemangioma.

In selected embodiments, the invention provides a method of treating asolid tumor cancer with a composition including a solid form of Formula(1) described herein, wherein the dose is effective to inhibit signalingbetween the solid tumor cells and at least one microenvironment selectedfrom the group consisting of macrophages, monocytes, mast cells, helperT cells, cytotoxic T cells, regulatory T cells, natural killer cells,myeloid-derived suppressor cells, regulatory B cells, neutrophils,dendritic cells, and fibroblasts. In selected embodiments, the inventionprovides a method of treating pancreatic cancer, breast cancer, ovariancancer, melanoma, lung cancer, squamous cell carcinoma including headand neck cancer, and colorectal cancer using a solid form of Formula (1)described herein, wherein the dose is effective to inhibit signalingbetween the solid tumor cells and at least one microenvironment selectedfrom the group consisting of macrophages, monocytes, mast cells, helperT cells, cytotoxic T cells, regulatory T cells, natural killer cells,myeloid-derived suppressor cells, regulatory B cells, neutrophils,dendritic cells, and fibroblasts. In an embodiment, the inventionprovides a method for treating pancreatic cancer, breast cancer, ovariancancer, melanoma, lung cancer, head and neck cancer, and colorectalcancer using a combination of a solid form of Formula (1) describedherein and a second agent selected from the group consisting ofbendamustine, venetoclax, gemcitabine, albumin-bound paclitaxel,rituximab, obinutuzumab, ofatumumab, pembrolizumab, nivolumab,durvalumab, avelumab, and atezolizumab, wherein the BTK inhibitor is asolid form of Formula (1) described herein. In an embodiment, the solidform of Formula (1) in any of the foregoing embodiments is Form I of thefree base.

In some embodiments, the invention relates to a method of treating aninflammatory, immune, or autoimmune disorder in a mammal with acomposition including a solid form of Formula (1) described herein. Inselected embodiments, the invention also relates to a method of treatinga disease with a composition including a solid form of Formula (1)described herein, wherein the disease is selected from the groupconsisting of tumor angiogenesis, chronic inflammatory disease,rheumatoid arthritis, atherosclerosis, inflammatory bowel disease, skindiseases such as psoriasis, eczema, and scleroderma, diabetes, diabeticretinopathy, retinopathy of prematurity, age-related maculardegeneration, hemangioma, glioma and melanoma, ulcerative colitis,atopic dermatitis, pouchitis, spondylarthritis, uveitis, Behcetsdisease, polymyalgia rheumatica, giant-cell arteritis, sarcoidosis,Kawasaki disease, juvenile idiopathic arthritis, hidratenitissuppurativa, Sjögren's syndrome, psoriatic arthritis, juvenilerheumatoid arthritis, ankylosing spoldylitis, Crohn's Disease, lupus,and lupus nephritis.

In some embodiments, the invention relates to a method of treating ahyperproliferative disorder in a mammal with a composition including asolid form of Formula (1) described herein, wherein thehyperproliferative disorder is a B cell hematological malignancyselected from the group consisting of chronic lymphocytic leukemia(CLL), small lymphocytic leukemia (SLL), non-Hodgkin's lymphoma (NHL),diffuse large B cell lymphoma (DLBCL), follicular lymphoma (FL), mantlecell lymphoma (MCL), Hodgkin's lymphoma, B cell acute lymphoblasticleukemia (B-ALL), Burkitt's lymphoma, Waldenström's macroglobulinemia(WM), Burkitt's lymphoma, multiple myeloma, myelodysplatic syndromes, ormyelofibrosis. In some embodiments, the invention relates to a method oftreating a hyperproliferative disorder in a mammal with a compositionincluding a solid form of Formula (1) described herein, wherein thehyperproliferative disorder is selected from the group consisting ofchronic myelocytic leukemia, acute myeloid leukemia, DLBCL (includingactivated B-cell (ABC) and germinal center B-cell (GCB) subtypes),follicle center lymphoma, Hodgkin's disease, multiple myeloma, indolentnon-Hodgkin's lymphoma, and mature B-cell ALL.

In some embodiments, the hyperproliferative disorder is a subtype ofCLL. A number of subtypes of CLL have been characterized. CLL is oftenclassified for immunoglobulin heavy-chain variable-region (IgV_(H))mutational status in leukemic cells. R. N. Damle, et al., Blood 1999,94, 1840-47; T. J. Hamblin, et al., Blood 1999, 94, 1848-54. Patientswith IgV_(H) mutations generally survive longer than patients withoutIgV_(H) mutations. ZAP70 expression (positive or negative) is also usedto characterize CLL. L. Z. Rassenti, et al., N. Engl. J. Med. 2004, 351,893-901. The methylation of ZAP-70 at CpG3 is also used to characterizeCLL, for example by pyrosequencing. R. Claus, et al., J. Clin. Oncol.2012, 30, 2483-91; J. A. Woyach, et al., Blood 2014, 123, 1810-17. CLLis also classified by stage of disease under the Binet or Rai criteria.J. L. Binet, et al., Cancer 1977, 40, 855-64; K. R. Rai, T. Han,Hematol. Oncol. Clin. North Am. 1990, 4, 447-56. Other common mutations,such as 11q deletion, 13q deletion, and 17p deletion can be assessedusing well-known techniques such as fluorescence in situ hybridization(FISH). In an embodiment, the invention relates to a method of treatinga CLL in a human, wherein the CLL is selected from the group consistingof IgV_(H) mutation negative CLL, ZAP-70 positive CLL, ZAP-70 methylatedat CpG3 CLL, CD38 positive CLL, chronic lymphocytic leukemiacharacterized by a 17p13.1 (17p) deletion, and CLL characterized by a11q22.3 (11q) deletion.

In some embodiments, the hyperproliferative disorder is a CLL whereinthe CLL has undergone a Richter's transformation. Methods of assessingRichter's transformation, which is also known as Richter's syndrome, aredescribed in P. Jain and S. O'Brien, Oncology, 2012, 26, 1146-52.Richter's transformation is a subtype of CLL that is observed in 5-10%of patients. It involves the development of aggressive lymphoma from CLLand has a generally poor prognosis.

In some embodiments, the hyperproliferative disorder is a CLL or SLL ina patient, wherein the patient is sensitive to lymphocytosis. In anembodiment, the invention relates to a method of treating CLL or SLL ina patient, wherein the patient exhibits lymphocytosis caused by adisorder selected from the group consisting of a viral infection, abacterial infection, a protozoal infection, or a post-splenectomy state.In an embodiment, the viral infection in any of the foregoingembodiments is selected from the group consisting of infectiousmononucleosis, hepatitis, and cytomegalovirus. In an embodiment, thebacterial infection in any of the foregoing embodiments is selected fromthe group consisting of pertussis, tuberculosis, and brucellosis.

Efficacy of the compounds and combinations of compounds described hereinin treating, preventing and/or managing other indicated diseases ordisorders described here can also be tested using various models knownin the art. Efficacy in treating, preventing and/or managing asthma canbe assessed using the ova induced asthma model described, for example,in Lee, et al., J. Allergy Clin. Immunol. 2006, 118, 403-9. Efficacy intreating, preventing and/or managing arthritis (e.g., rheumatoid orpsoriatic arthritis) can be assessed using the autoimmune animal modelsdescribed in, for example, Williams, et al., Chem. Biol. 2010, 17,123-34, WO 2009/088986, WO 2009/088880, and WO 2011/008302. Efficacy intreating, preventing and/or managing psoriasis can be assessed usingtransgenic or knockout mouse model with targeted mutations in epidermis,vasculature or immune cells, mouse model resulting from spontaneousmutations, and immuno-deficient mouse model with xenotransplantation ofhuman skin or immune cells, all of which are described, for example, inBoehncke, et al., Clinics in Dermatology, 2007, 25, 596-605. Efficacy intreating, preventing and/or managing fibrosis or fibrotic conditions canbe assessed using the unilateral ureteral obstruction model of renalfibrosis, which is described, for example, in Chevalier, et al., KidneyInternational 2009, 75, 1145-1152; the bleomycin induced model ofpulmonary fibrosis described in, for example, Moore, et al., Am. J.Physiol. Lung. Cell. Mol. Physiol. 2008, 294, L152-L160; a variety ofliver/biliary fibrosis models described in, for example, Chuang, et al.,Clin. Liver Dis. 2008, 12, 333-347 and Omenetti, et al., LaboratoryInvestigation, 2007, 87, 499-514 (biliary duct-ligated model); or any ofa number of myelofibrosis mouse models such as described in Varicchio,et al., Expert Rev. Hematol. 2009, 2, 315-334. Efficacy in treating,preventing and/or managing scleroderma can be assessed using a mousemodel induced by repeated local injections of bleomycin described, forexample, in Yamamoto, et al., J. Invest. Dermatol. 1999, 112, 456-462.Efficacy in treating, preventing and/or managing dermatomyositis can beassessed using a myositis mouse model induced by immunization withrabbit myosin as described, for example, in Phyanagi, et al., Arthritis& Rheumatism, 2009, 60(10), 3118-3127. Efficacy in treating, preventingand/or managing lupus can be assessed using various animal modelsdescribed, for example, in Ghoreishi, et al., Lupus, 2009, 19,1029-1035; Ohl, et al., J. Biomed. & Biotechnol., Article ID 432595(2011); Xia, et al., Rheumatology, 2011, 50, 2187-2196; Pau, et al.,PLoS ONE, 2012, 7(5), e36761; Mustafa, et al., Toxicology, 2011, 90,156-168; Ichikawa et al., Arthritis & Rheumatism, 2012, 62(2), 493-503;Rankin, et al., J. Immunology, 2012, 188, 1656-1667. Efficacy intreating, preventing and/or managing Sjögren's syndrome can be assessedusing various mouse models described, for example, in Chiorini, et al.,J. Autoimmunity, 2009, 33, 190-196.

Efficacy of the compounds and combinations of compounds described hereinin treating, preventing and/or managing the indicated diseases ordisorders can be tested using various models known in the art. Forexample, models for determining efficacy of treatments for pancreaticcancer are described in Herreros-Villanueva, et al., World J.Gastroenterol. 2012, 18, 1286-1294. Models for determining efficacy oftreatments for breast cancer are described, e.g., in Fantozzi, BreastCancer Res. 2006, 8, 212. Models for determining efficacy of treatmentsfor ovarian cancer are described, e.g., in Mullany, et al.,Endocrinology 2012, 153, 1585-92; and Fong, et al., J. Ovarian Res.2009, 2, 12. Models for determining efficacy of treatments for melanomaare described, e.g., in Damsky, et al., Pigment Cell & Melanoma Res.2010, 23, 853-859. Models for determining efficacy of treatments forlung cancer are described, e.g., in Meuwissen, et al., Genes &Development, 2005, 19, 643-664. Models for determining efficacy oftreatments for lung cancer are described, e.g., in Kim, Clin. Exp.Otorhinolaryngol. 2009, 2, 55-60; and Sano, Head Neck Oncol. 2009, 1,32. Models for determining efficacy of treatments for colorectal cancer,including the CT26 model, are described in Castle, et al., BMC Genomics,2013, 15, 190; Endo, et al., Cancer Gene Therapy, 2002, 9, 142-148; Rothet al., Adv. Immunol. 1994, 57, 281-351; Fearon, et al., Cancer Res.1988, 48, 2975-2980.

Models for determining efficacy of treatments in hematologicalmalignancies, including B cell cancers, may also be used. For example,efficacy in diffuse large B cell lymphoma (DLBCL) may be assessed usingthe PiBCL1 murine model and BALB/c (haplotype H-2d) mice. Illidge, etal., Cancer Biother. & Radiopharm. 2000, 15, 571-80. Efficacy innon-Hodgkin's lymphoma (NHL) may be assessed using the 38C13 murinemodel with C3H/HeN (haplotype 2-Hk) mice or alternatively the 38C13Her2/neu model. Timmerman, et al., Blood 2001, 97, 1370-77; Penichet, etal., Cancer Immunolog. Immunother. 2000, 49, 649-662. Efficacy in CLLmay be assessed using the BCL1 model using BALB/c (haplotype H-2d) mice.Dutt, et al., Blood, 2011, 117, 3230-29.

EXAMPLES Example 1 Form I of(S)-4-(8-Amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide(Free Base) Crystalline Anhydrate Example 1.1 Preparation of Form ICrystalline Anhydrate

A crystallization study was performed using amorphous(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideas input. The amorphous character of this batch was confirmed by PXRD.For cooling crystallization experiments, 25 mg of amorphous Formula (1)was dissolved in 300 μL solvent, heated to 60° C. at a rate of 5°C./hour, held for 1 hour at that temperature, and then cooled down to 5°C. at the same rate. For slurry experiments, 25 mg of amorphous Formula(1) was suspended in 150 μL solvent at 20° C. for 3 days. All solidswere isolated for PXRD analysis. The solvents were evaporated undervacuum (200 mbar) when a clear solution was obtained. The results aresummarized in Table 1. The results indicate that, when solids areobtained, the amorphous form of Formula (1) is obtained from mostsolvents, and that Form I is difficult to crystallize but may beprepared from a very limited set of solvents, in particular certainmixtures with n-heptane (e.g., with acetone). Form I may also becrystallized or recrystallized from ethanol at larger scales, includingat 60 g scale.

TABLE 1 Results of crystallization and slurry experiments for(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-yl)-N-(pyridin-2-yl)benzamide.Appearance after Sample Solvent Type Appearance evaporation PXRD DSC TGA1 methanol Cooling crystallization Dissolved Solid Amorphous — — 2ethanol Cooling crystallization Dissolved Gel — — — 3 2-propanol Coolingcrystallization Gel Gel — — — 4 N,N-dimethylacetamide Coolingcrystallization Dissolved Gel — — — 5 acetone Cooling crystallizationDissolved Solid Amorphous — — 6 2-butanone Cooling crystallizationDissolved Gel — — — 7 cyclohexanone Cooling crystallization DissolvedGel — — — 8 dimethyl sulfoxide Cooling crystallization Dissolved Gel — —— 9 chlorobenzene Cooling crystallization Solid — Amorphous — — 10dichloromethane Cooling crystallization Dissolved Solid Amorphous — — 11methanol-water 3:1 Cooling crystallization Dissolved Solid Amorphous — —12 methanol-water 1:1 Cooling crystallization Gel Gel — — — 13methanol-water 1:3 Cooling crystallization Gel Gel — — — 14ethanol-water 3:1 Cooling crystallization Dissolved Solid Amorphous — —15 ethanol-water 1:1 Cooling crystallization Gel Gel — — — 16ethanol-water 1:3 Cooling crystallization Gel Gel — — — 172-propanol-water 3:1 Cooling crystallization Dissolved Solid Amorphous —— 18 2-propanol-water 1:1 Cooling crystallization Dissolved SolidAmorphous — — 19 2-propanol-water 1:3 Cooling crystallization Gel Gel —— — 20 N,N-dimethylacetamide-water 3:1 Cooling crystallization DissolvedGel — — — 21 N,N-dimethylacetamide-water 1:1 Cooling crystallizationDissolved Gel — — — 22 N,N-dimethylacetamide-water 1:3 Coolingcrystallization Gel Gel — — — 23 acetone-heptane 3:1 Coolingcrystallization Solid — Form I — — 24 acetone-heptane 1:1 Coolingcrystallization Solid — Form I — — 25 acetone-heptane 1:3 Coolingcrystallization Gel Gel — — — 26 2-butanone-heptane 3:1 Coolingcrystallization Gel Gel — — — 27 2-butanone-heptane 1:1 Coolingcrystallization Gel Gel — — — 28 2-butanone-heptane 1:3 Coolingcrystallization Solid — Amorphous — — 29 cyclohexanone-heptane 3:1Cooling crystallization Dissolved Gel — — — 30 cyclohexanone-heptane 1:1Cooling crystallization Gel Gel — — — 31 cyclohexanone-heptane 1:3Cooling crystallization Gel Gel — — — 32 dimethyl sulfoxide-water 3:1Cooling crystallization Gel Gel — — — 33 dimethyl sulfoxide-water 1:1Cooling crystallization Gel Gel — — — 34 dimethyl sulfoxide-water 1:3Cooling crystallization Solid — Amorphous — — 35 chlorobenzene-heptane3:1 Cooling crystallization Solid — Amorphous — — 36chlorobenzene-heptane 1:1 Cooling crystallization Solid — Amorphous — —37 chlorobenzene-heptane 1:3 Cooling crystallization Solid — Amorphous —— 38 dichloromethane-heptane 3:1 Cooling crystallization Dissolved SolidAmorphous — — 39 dichloromethane-heptane 1:1 Cooling crystallizationSolid — Form I 207° C. −1.6% (−161 J/g) (40-140° C.) −1.2% (150-240° C.)40 dichloromethane-heptane 1:3 Cooling crystallization Gel Gel — — — 41methyl tert-butyl ether Slurry (20° C. for 3 days) Solid — Amorphous — —42 tetrahydrofuran Slurry (20° C. for 3 days) Dissolved Solid Amorphous— — 43 diisopropyl ether Slurry (20° C. for 3 days) Solid — Amorphous —— 44 2-methyltetrahydrofuran Slurry (20° C. for 3 days) Solid —Amorphous — — 45 cyclopentyl methyl ether Slurry (20° C. for 3 days)Solid — Amorphous — — 46 methanol-water 3:1 Slurry (20° C. for 3 days)Dissolved Solid Amorphous — — 47 methanol-water 1:1 Slurry (20° C. for 3days) Gel Gel — — — 48 methanol-water 1:3 Slurry (20° C. for 3 days) GelGel — — — 49 ethanol-water 3:1 Slurry (20° C. for 3 days) DissolvedSolid Amorphous — — 50 ethanol-water 1:1 Slurry (20° C. for 3 days) GelGel — — — 51 ethanol-water 1:3 Slurry (20° C. for 3 days) Gel Gel — — —52 2-propanol-water 3:1 Slurry (20° C. for 3 days) Dissolved SolidAmorphous — — 53 2-propanol-water 1:1 Slurry (20° C. for 3 days) Solid —Form II ~105° C. −9.7% ~150° C. (40-120° C.) ~220° C. 542-propanol-water 1:3 Slurry (20° C. for 3 days) Gel Gel — — — 55N,N-dimethylacetamide-water 3:1 Slurry (20° C. for 3 days) Dissolved Gel— — — 56 N,N-dimethylacetamide-water 1:1 Slurry (20° C. for 3 days)Dissolved Gel — — — 57 N,N-dimethylacetamide-water 1:3 Slurry (20° C.for 3 days) Gel Gel — — — 58 acetone-heptane 3:1 Slurry (20° C. for 3days) Solid — Form I — — 59 acetone-heptane 1:1 Slurry (20° C. for 3days) Gel Gel — — — 60 acetone-heptane 1:3 Slurry (20° C. for 3 days)Solid — Amorphous — — 61 2-butanone-heptane 3:1 Slurry (20° C. for 3days) Gel Gel — — — 62 2-butanone-heptane 1:1 Slurry (20° C. for 3 days)Solid — Amorphous — — 63 2-butanone-heptane 1:3 Slurry (20° C. for 3days) Solid — Amorphous — — 64 cyclohexanone-heptane 3:1 Slurry (20° C.for 3 days) Dissolved Gel — — — 65 cyclohexanone-heptane 1:1 Slurry (20°C. for 3 days) Gel Gel — — — 66 cyclohexanone-heptane 1:3 Slurry (20° C.for 3 days) Solid — Amorphous — — 67 dimethyl sulfoxide-water 3:1 Slurry(20° C. for 3 days) Solid — Poorly — — crystalline 68 dimethylsulfoxide-water 1:1 Slurry (20° C. for 3 days) Solid — Amorphous — — 69dimethyl sulfoxide-water 1:3 Slurry (20° C. for 3 days) Solid — Form II— — 70 chlorobenzene-heptane 3:1 Slurry (20° C. for 3 days) Solid —Amorphous — — 71 chlorobenzene-heptane 1:1 Slurry (20° C. for 3 days)Solid — Amorphous — — 72 chlorobenzene-heptane 1:3 Slurry (20° C. for 3days) Solid — Amorphous — — 73 dichloromethane-heptane 3:1 Slurry (20°C. for 3 days) Dissolved Solid Amorphous — — 74 dichloromethane-heptane1:1 Slurry (20° C. for 3 days) Gel Gel — — — 75 dichloromethane-heptane1:3 Slurry (20° C. for 3 days) Solid — Amorphous — —

Anti-solvent addition experiments were performed by stepwise addition ofanti-solvent until crystallization, to a clear solution of Formula (1)in the solvent shown in Table 2. The results again highlight thedifficulty in preparing crystalline Formula (1).

TABLE 2 Results of anti-solvent addition experiments for(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide.Sample Solvent Type Anti-solvent Appearance¹ PXRD 76 methanolAnti-solvent Water FFP Amorphous 77 ethanol Anti-solvent Water FFPAmorphous 78 2-propanol Anti-solvent Water No solids — 79 N,N-Anti-solvent Water FFP Amorphous 80 acetone Anti-solvent Heptane Stickysolids Amorphous 81 2-butanone Anti-solvent Heptane Sticky solidsAmorphous 82 cyclohexanone Anti-solvent Heptane FFP Amorphous 83dimethyl sulfoxide Anti-solvent Water FFP + sticky solids Amorphous 84chlorobenzene Anti-solvent Heptane FFP + sticky solids Amorphous 85dichloromethane Anti-solvent Heptane FFP Amorphous ¹FFP refers to freeflowing powder.

Example 1.2 Physical Characterization of Form I Crystalline Anhydrate

Characterization of Form I of the free base of Formula (1) produced bycrystallization from acetone in the presence of methanol (referred to assample PP502-P1 herein) was carried out using various techniquesincluding: PXRD (FIG. 1 and FIG. 2), optical microscopy (FIG. 3), Raman(FIG. 4) and IR spectroscopy (FIG. 5 and FIG. 6), solution-state NMRspectroscopy after dissolution of Form I (FIG. 7), TG-FTIR (FIG. 8),differential scanning calorimetry (DSC) (FIG. 9), semi-quantitativesolubility testing, and dynamic vapor sorption (DVS; also known asgravimetric vapor sorption or GVS) (FIGS. 10 and 11).

The transmission PXRD pattern of Form I was acquired using a Stoe StadiP high-precision two circle goniometer instrument equipped with a Mythen1K Detector and a Cu-K_(α1) radiation source operating at standardmeasurement conditions of: 40 kV tube voltage and 40 mA tube current;curved Ge monochromator; 0.02 °2θ step size; 48 seconds step time,1.5-50.5 °2θ scanning range; and detector mode including a step scan at1 °2θ detector step. Samples were prepared by placing 10 to 20 mg ofmaterial between two acetate foils in the Stoe transmission sampleholder which was rotated during measurement. The measurements using theStoe Stadi diffractometer were taken in transmission (Debye-Scherrer)mode. This instrument can also be operated in reflection(Bragg-Brentano) mode.

Reflection PXRD measurements were also performed using a secondinstrument, a Bruker D8 Advance powder X-ray diffractometer equippedwith a LynxEye detector and operating in Bragg-Brentano reflectiongeometry. 2θ values are generally accurate to within an error of ±0.2°.The samples were generally prepared without any special treatment otherthan the application of slight pressure to get a flat surface. Sampleswere measured uncovered unless otherwise noted. Operating conditionsincluded a tube voltage of 40 kV and current of 40 mA. A variabledivergence slit was used with a 3° window. The step size was 0.02 °2θwith a step time of 37 seconds. The sample was rotated at 0.5 rps duringthe measurement. When calibrated, the reflection mode PXRD pattern ofForm I may be compared to the transmission mode PXRD pattern of Form I,although the person of skill in the art will appreciate that thediffraction patterns may vary, particularly with respect to peakintensities, as described herein.

FIG. 1 shows the PXRD pattern for Form I of Formula (1) measured usingreflection geometry. The following peaks were identified in the PXRDpattern of FIGS. 1: 6.4, 8.6, 10.5, 10.9, 11.3, 11.6, 12.7, 13.4, 14.3,14.9, 15.1, 15.7, 16.1, 17.3, 18.2, 19.2, 19.4, 19.8, 20.7, 21.1, 21.4,21.6, 21.9, 22.6, 23.3, 23.6, 24.9, 25.2, 25.4, 25.7, 26.1, 26.4, 26.8,26.9, 27.7, 28.6, 29.1, 29.4, 30.1, 30.5, 31.7, 31.9, 32.2, 32.6, 33.1,33.4, 34.5, 35.9, 36.1, 36.8, 37.4, 38.1, 38.9, and 39.5 °2θ±0.2 °2θ.

FIG. 2 shows the PXRD pattern for Form I measured using transmissiongeometry. The following peaks were identified in the PXRD pattern ofFIGS. 2: 6.4, 8.7, 10.5, 11.0, 11.4, 11.6, 12.8, 13.5, 14.3, 14.9, 15.1,15.5, 15.7, 16.1, 17.3, 18.2, 19.1, 19.2, 19.5, 19.8, 20.6, 20.8, 21.2,21.4, 21.6, 22.0, 22.2, 22.3, 22.6, 22.8, 23.3, 23.7, 24.9, 25.2, 25.4,25.8, 26.1, 26.5, 26.8, 27.0, 27.0, 27.7, 28.7, 29.2, 29.9, 30.5, 31.7,32.0, 32.6, 33.1, 33.2, 33.5, 34.5, and 35.1 °2θ±0.2 °2θ. Form I showsdistinctive peaks (relative to the other forms) at 6.4, 8.6, 10.5, 11.6,and 15.7 °2θ±0.2 °2θ, and shows further distinctive peaks (relative tothe other forms) at 10.9, 12.7, 13.4, 14.3, 14.9, and 18.2 °2θ±0.2 °2θ.

Both the PXRD patterns of FIG. 1 and FIG. 2, along with thebirefringence observed in the polarized optical microscopy images ofFIG. 3, show that the anhydrate of Form I of Formula (1) is crystalline.

The Fourier-transform (FT) Raman spectrum of Form I was acquired using aBruker RFS 100 FT-Raman spectrophotometer equipped with a liquidnitrogen-cooled germanium detector and a near IR Nd:YAG laser operatingat 1064 nm with a power setting of 100 mW. Spectra were the result of 64scans collected with a resolution of 2 cm⁻¹ in the range between 3500and 50 cm⁻¹. The FT-Raman spectrum of Form I in the relevant fingerprintregion is shown in FIG. 4, and has peaks at 1680, 1620, 1609, 1574,1547, 1514, 1495, 1454, 1433, 1351, 1312, 1255, 1232, 1187, 1046, 995,706, 406, and 280 (Raman shift, cm⁻¹±2 cm⁻¹).

The IR spectrum of Form I (sample PP502-P1) was obtained using IRspectroscopy.

The spectra were obtained by recording 32 scans using attenuated totalreflectance (ATR) sampling and a Perkin Elmer BXII IR spectrometer at aresolution of 2 wavenumbers (cm⁻¹). For the spectrum shown here, theoriginal spectra in transmission mode were converted to absorption modeusing the OPUS 7.0 software from Bruker and peak tables were generated.The IR spectrum of Form I is illustrated in FIG. 5, and an expansion ofthe region from 1800 to 600 cm⁻¹ is shown in FIG. 6. The characteristicpeaks for Form I are given in Table 3.

TABLE 3 Characteristic peaks for Form I as determined by ATR IRspectroscopy (+/−4 cm⁻¹). Wavenumber Intensity (cm⁻¹) (arbitrary units)3367 0.088 3089 0.098 2246 0.071 1682 0.208 1621 0.319 1608 0.371 15740.212 1514 0.304 1504 0.278 1454 0.141 1428 0.322 1403 0.493 1345 0.1521303 0.340 1248 0.181 1194 0.156 1177 0.156 1149 0.153 1109 0.119 10490.139 1023 0.162 1003 0.236 947 0.116 900 0.145 858 0.233 842 0.213 8160.221 764 0.345 734 0.295 729 0.290 701 0.234 689 0.186 665 0.134 6230.181 612 0.195

The ¹H NMR spectrum of Form I recorded in deuterated dimethyl sulfoxide(d₆-DMSO) is illustrated in FIG. 7 and confirms the molecular structureof(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidepresent in this crystalline anhydrate.

TGA and TG-FTIR analysis was carried out using a NetzschThermo-Microbalance TG 209 coupled to a Bruker FTIR spectrometer(instrument model Vector 22). The sample pans having a pinhole weretared before the sample was introduced and then and heated to 350° C. ata ramp rate of 10° C./minute under a constant flow of a nitrogen.TG-FTIR analysis of the sample of Form I revealed a mass loss of about0.8% upon heating to 250° C. TG-FTIR spectroscopy showed that theobserved mass loss up to 250° C. is essentially attributable to acetonesolvent, which appears to be tightly bound to the Form I crystal sincethe mass loss occurs above 200° C. Weight loss above 250° C. is mostlyattributable to decomposition. The TGA thermogram obtained from theTG-FTIR experiment is depicted in FIG. 8.

Differential scanning calorimetry was carried out with a Perkin ElmerDSC-7 or with a TA Instruments Q2000 instrument. Samples were preparedin a closed gold sample pan at temperature ramp rates of 10° C./minuteor 20° C./minute up to approximately 250° C. Melting begins at about200° C. and a peak is observed near 215° C. with a heat flow ofapproximately 16 mW for the melting endotherm; however, it appears thatmelting is concurrent with thermal decomposition and the enthalpy offusion cannot be evaluated. Nevertheless, the temperature range of themass loss observed in the TGA analysis of FIG. 8 suggests that Form Imust be molten in order to release the residual solvent. The DSCthermogram is shown in FIG. 9, where endothermic events are plotted inthe upward direction. After the melting event, exothermic degradationoccurs at 226.4° C.

Form I was also tested with respect to solubilities in variouswater-solvent mixtures and non-aqueous solvents. Solubility studies wereconducted by a stepwise dilution of a suspension of about 10 mg of FormI in 0.1 mL of analytical grade solvent. Results of the approximatesolubilities are shown in Table 4. Solubility values are estimatedapproximations and are subject to variable experimental error.

TABLE 4 Approximate solubility measurements for Form I. Solubility [S,Solubility [S, Pure Solvent mg/mL] Solvent mixture mg/mL] acetic acid102 < S < 204 acetic acid:water 1:1 104 < S < 208 acetone S~2 aceticacid:ethyl acetate 1:1 92 < S < 184 acetonitrile S < 1 acetic acid:ethylacetate 1:9 60 < S < 90 dichloromethane, DCM 36 < S < 43 acetic acid:MEK1:9 63 < S < 95 N,N-dimethylformamide, 49 < S < 65 aceticacid:isopropanol 1:9 S~4 DMF dimethyl sulfoxide, DMSO 39 < S < 49acetone:water 4:1 26 < S < 31 ethyl acetate S < 1 ethanol:water 1:1 S~6ethanol S~3 ethanol:water 9:1 at 60° C. S > 60 formic acid 97 < S < 194ethanol:water 95:5 S~8 2-butanone, MEK S < 1 MEK saturated with water 26< S < 30 methanol S~14 methanol:MEK 1:1 at reflux S > 90N-methyl-2-pyrrolidone, 39 < S < 49 methanol:water 9:1 S~15 NMP2-propanol S < 1 THF:water 9:1 S > 50 tetrahydrofuran, THF S~5trifluoroethane 97 < S < 194

The aqueous solubility of Form I was determined after equilibrating at25° C. for three days. High-performance liquid chromatography (HPLC) wasused to determine the concentration in filtered solution, which resultedin S˜68 μg/mL. PXRD of the solid residue confirmed that Form I wasretained.

A gravimetric vapor sorption study was run using a standard procedure.Samples were run using a dynamic vapor sorption (DVS) analyzer. Samplesizes were approximately 10 mg. A moisture adsorption-desorptionisotherm was performed as outlined below. Samples were exposed to astarting 50% RH, decreasing humidity to 0% RH, increasing humidity to95% RH, and finally decreasing humidity back to the starting 50% RH. TheDVS results, including sorption and desorption isotherm curves, shown inFIG. 10 and FIG. 11, show the total weight gain observed between 0% RHand 80% RH to be about 0.17%, which indicates that Form I isnon-hygroscopic according to the European Pharmacopoeia (EP)classification (non-hygroscopic: <0.2%; slightly hygroscopic: ≧0.2% and<2%; hygroscopic: ≧2% and <15%; very hygroscopic: ≧15%; deliquescent:sufficient water is absorbed to form a liquid; all values measured asweight increase at 80% RH and 25° C.). The desorption curve indicatesthat Form I lost moisture at a similar rate to the moisture gainedduring sorption, with limited hysteresis.

Almost all of the adsorbed water was removed by the end of the DVSexperiment. No form change was observed by PXRD after the DVSexperiment.

Example 2 Form II of(S)-4-(8-Amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide(Free Base) Trihydrate Example 2.1 Preparation of Form II CrystallineTrihydrate

The crystallization study described above and reported in Table 1 alsoyielded Form II in a very limited set of solvents.

Form II (sample PP502-P21) of the free base of Formula (1) was alsoproduced by dissolving Form I in an acetone-water (8:2) mixture atreflux temperature followed by cooling of the solution and removing 50%of the solvent volume under a slight nitrogen purge. The obtainedsamples were dried at room temperature in air and under ambientconditions (at about 45% RH). A mass loss of about 9.7% was observedafter drying, corresponding to approximately 2.7 water molecules permolecule of Formula (1) (i.e., a trihydrate).

Example 2.2 Physical Characterization of Form II Crystalline Trihydrate

Characterization of Form II of the free base of Formula (1) was carriedout using various techniques including PXRD (FIG. 12), opticalmicroscopy (FIG. 13), Raman spectroscopy (FIG. 14), IR spectroscopy(FIG. 15 and FIG. 16), TG-FTIR (FIG. 17), DSC (FIG. 18), DVS (FIG. 19and FIG. 20), and semi-quantitative solubility testing. Characterizationmethods used for Form II were performed as described previously for thecharacterization of Form I.

FIG. 12 shows the PXRD pattern for Form II of Formula (1) measured intransmission mode. The following characteristic peaks were identified inthe PXRD pattern of FIG. 12: 6.6, 9.9, 11.0, 13.6, 14.0, 14.3, 18.1,18.4, 18.9, 19.3, 20.2, 21.1, 22.0, 22.2, 22.5, 22.7, 22.9, 23.4, 23.5,23.9, 24.2, 24.6, 25.0, 26.1, 26.6, 26.9, 27.5, 28.2, 31.0, 32.1, 32.4,32.7, 33.4, 33.9, and 34.4 °2θ±0.2 °2θ. An optical microscopic image ofForm II in FIG. 13 shows that the Form II sample (PP502-P21) exhibitsrod-shaped particles with lengths up to about 50 μm.

The FT-Raman spectrum of Form II in the relevant fingerprint region (200cm⁻¹ to 1800 cm⁻¹) is shown in FIG. 14 and exhibits peaks (Raman shift,cm⁻¹±2 cm⁻¹) at 1668, 1611, 1580, 1564, 1537, 1506, 1493, 1454, 1436,1416, 1401, 1349, 1321, 1287, 1272, 1252, 1244, 1183, 1165, 1097, 1039,1025, 996, 950, 871, 853, 776, 730, 645, 633, 375, 352, 279, and 247.

FIG. 15 shows the IR spectrum of Form II (sample PP502-P21), and anexpansion of the region from 1800 to 600 cm⁻¹ is shown in FIG. 16. Thecharacteristic peaks observed for Form II are provided in Table 5.

TABLE 5 Characteristic peaks for Form II as determined by ATR IRspectroscopy (+/−4 cm⁻¹). Wavenumber Intensity (cm⁻¹) (arbitrary units)3212 0.146 2206 0.061 1665 0.225 1618 0.449 1577 0.370 1548 0.339 15350.283 1504 0.332 1465 0.155 1452 0.153 1432 0.515 1416 0.404 1397 0.4061348 0.169 1316 0.529 1243 0.181 1208 0.178 1181 0.122 1164 0.141 11490.177 1095 0.129 1038 0.139 1004 0.194 948 0.144 891 0.204 869 0.201 8210.257 776 0.436 736 0.393 716 0.432 643 0.250 617 0.346

TG-FTIR preparation of Form II samples consisted of exposing two samples(PP502-P14 and PP502-P21) to 60% RH for about three days at which pointboth contained identical amounts of water. TG-FTIR analysis of thesamples of Form II, shown in FIG. 17, revealed a mass loss of about10.2% upon heating to approximately 130° C. This decrease is essentiallyattributable to release of water and agrees well with the theoreticalwater content for a trihydrate of 10.4%. Mass loss of about 0.3% uponheating thereafter to approximately 250° C. was due primarily todecomposition.

A representative DSC analysis of a Form II sample is shown in FIG. 18.The sample was stabilized at equilibrium under approximately 62% RHbefore analysis using temperature ramp rates of 10° C./minute or 20°C./minute up to approximately 150° C. Melting begins at about 75° C. anda peak is observed near about 109° C. with an enthalpy of fusion ofabout 127 J/g. The DSC thermogram shows a slight shoulder on the leftside of the peak that suggests that some of the hydrate water might havebeen released from the sample into the residual volume of thehermetically sealed sample pan.

DVS analysis of the Form II (sample PP502-P14) was performed by exposingsamples to a starting 50% RH, decreasing humidity to 0% RH, increasinghumidity to 95% RH, and finally decreasing humidity back to the starting50% RH. The DVS results, including sorption and desorption curves, areshown in FIG. 19 and FIG. 20. The results show that significant waterloss occurs below about 10% RH, when the trihydrate water contentrapidly decreases from approximately 10% to approximately 0%. Thisresult is consistent with the mass loss in the TGA analysis. Uponincreasing the RH to 95%, water was re-adsorbed to achieve a maximumwater content of about 10.4%, which corresponds to the expected watercontent for a trihydrate. Hysteresis was also observed between thesorption and desorption curves. Form II thus behaves as a variablehydrate. The DVS results, including sorption and desorption isothermcurves, show the total weight gain observed between 0% RH and 80% RH tobe about 10%, which indicates that Form II is hygroscopic according tothe EP classification (see Example 1.2

Finally, Form II was tested with respect to solubilities in variouswater-solvent mixtures and non-aqueous solvents. The aqueous solubilityof Form II was determined after equilibrating at 25° C. for three days.High-performance liquid chromatography (HPLC) was used to determine theconcentration of Form II in the filtered solution at approximately 14μg/mL, which translates into a critical water activity (a_(w)) of about0.59. In comparison, the aqueous solubility for Form I is about 68μg/mL.

Critical water activity is a measure of the relative thermodynamicstability of Form I in comparison to the trihydrate Form II. Below ana_(w) of about 0.59, Form I is more stable at room temperature whileabove this value Form II is more stable. Suspension equilibrationexperiments in ethanol-water mixtures, each having a different wateractivity, affirmed this conclusion. Water activities included in theexperiments were maintained at: about 0.35 (ethanol-water ratio of 95:5,PP502-P32), about 0.53 (ethanol-water ratio of 9:1, PP502-P33) and about0.77 (ethanol-water ratio of 7:3, PP502-P34). At an a_(w) of about 0.53,suspension experiments having mixtures of Form I and Form II resulted inpure Form I and at an a_(w) of about 0.77, the result was pure Form II.

Example 3 Form III of(S)-4-(8-Amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide(Free Base) Dihydrate Example 3.1 Preparation of Form III CrystallineDihydrate

Form III of the free base of Formula (1) was prepared from seededcrystallization experiments using Form I seeds. Saturated solutions ofFormula (1) were prepared at 60° C. The solutions were cooled and seedsof Form I were added before spontaneous crystallization occurred. Theresults are summarized in Table 6.

TABLE 6 Results of seeded crystallization experiments for(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide.Sample Solvent Type Appearance PXRD DSC TGA 86 methanol-water 1:1Seeding Solid Form III 147° C. (−23.7 J/g) −4.8% (40-130° C.) 215° C.(141 J/g)  87 ethanol-water 1:3 Seeding Solid Form III — — 882-propanol-water 1:3 Seeding Solid Form III — — 89N,N-dimethylacetamide-water 1:2 Seeding Solid Poorly — — crystalline 90Acetone-heptane 1:1 Seeding Solid Form I — — 91 2-butanone-heptane 1:1Seeding Gel — — — 92 cyclohexanone-heptane 1:1 Seeding Solid Form I — —93 dimethyl sulfoxide-water 1:3 Seeding Solid Poorly — — crystalline 94methyl tert-butyl ether Seeding Solid Amorphous — — 95tetrahydrofuran-water 1:6 Seeding Solid Form III — — 96 diisopropylether Seeding Solid Amorphous — — 97 2-methyltetrahydrofuran SeedingSolid Poorly — — crystalline 98 cyclopentyl methyl ether Seeding SolidPoorly — — crystalline 99 chlorobenzene Seeding Solid Poorly — —crystalline 100 dichloromethane-water 1:3 Seeding Gel — — —

Form III may be prepared by crystallization of amorphous Formula (1)free base in pure water. For instance, sample PP502-P120 was the resultsof a slurry of amorphous Formula (1) free base (sample PP502-P107A) inwater. After one day, Form III was found in the suspension, and after anextended stirring period of three days, Form III was still retained.However, because Form II may also be obtained in other experiments undersimilar conditions, additional procedures to prepare Form III weredevelopment.

Form III may also been prepared from amorphous Formula (1) suspended inwater. To about 160 mg of amorphous Formula (1), 5.0 mL of water isadded and the resulting suspension is stirred at ambient temperature.Investigation of the solid after about 24 hours of equilibration at roomtemperature led to crystallization of Form III.

Form III may also be prepared by direct precipitation via pH adjustment.940 mg of Formula (1) Form I is dissolved in 4.0 mL of 1 N aqueoushydrochloric acid solution. The solution neutralized with the sameamount of 1 N aqueous sodium hydroxide solution. Further dilution with8.0 mL of water leads to a thick suspension from which the solid isseparated by filtration. The glass bottle is rinsed with 16 mL of waterand the wash liquid is poured onto the glass frit filter and pulledthrough the filtration unit by application of vacuum. The obtained solidmaterial is dried in an air dryer at 40° C. for about 24 hours. PowderX-ray diffraction confirms that Formula (1) Form III is obtained andthermogravimetry coupled with infrared spectroscopy shows that thesample contains about 6% water, which suggests that the material wasslightly overdried. The water content found is still consistent with theresult from the DVS testing as this water content is found near 40% RH.

Example 3.2 Physical Characterization of Form III Crystalline Dihydrate

The PXRD, TGA, and DSC characterization methods used for Form III of thefree base of Formula (1) were performed as follows. The PXRD studieswere performed using a Bruker AXS D2 PHASER operating in theBragg-Brentano configuration. Data was collected using a Cu anode at 30kV and 10 mA with the sample rotating. Monochromatisation was performedusing a Kj3-filter (0.5% Ni). Fixed divergence slits were set at 1.0 mm(0.61°), the primary axial Soller slit was set at 2.5°, and thesecondary axial Soller slit was set at 2.5°. The detector was a linearLYNXEYE detector with a receiving slit 5° detector opening. The standardsample holder (0.1 mm cavity in (510) silicon wafer) has a minimalcontribution to the background signal. Measurement conditions were asfollows: scan range 5-45° 29, sample rotation 5 rpm, 0.5 s/step,0.010°/step, 3.0 mm detector slit; and all measuring conditions arelogged in the instrument control file. As a check of system suitability,a corundum sample A26-B26-S(NIST standard) is measured daily. Thesoftware used for PXRD data collection is Diffrac.Commander v3.3.35.Data analysis was performed using Diffrac.Eva V3.0 software. Nobackground correction or smoothing was applied to the patterns. Thecontribution of the Cu-Kα2 peak was stripped off using the Diffrac.Evasoftware. TGA and DSC studies were performed using a Mettler ToledoTGA/DSC1 STARe System with a 34-position auto sampler. The samples wereprepared using aluminium crucibles (40 μL; pierced). Typically, 5-10 mgof sample was loaded into a pre-weighed aluminium crucible and was keptat 30° C. for 5 minutes, after which it was heated at 10° C./min from30° C. to 300° C. A nitrogen purge of 40 mL/min was maintained over thesample. As system suitability check, indium and zinc are used asreferences. The software used for data collection and evaluation isSTARe Software v10.00 build 2480. No corrections are applied to thethermogram.

Additional PXRD characterization of Form III was obtained in a similarmanner as described in Example 1.2 for Form I. This PXRD pattern, whichwas obtained from sample PP502-P120 (prepared as described above), isshown in FIG. 21. The following peaks were identified in the PXRDpattern of FIG. 6: 10.4, 12.6, 12.8, 17.9, 21.3, 21.7, 23.1, 24.2, 25.2,and 27.0 °2θ±0.2 °2θ. Form III shows distinctive peaks (relative to theother forms) at 7.6, 8.5, 12.6, 12.8, 14.6, 16.8, and 23.2 °2θ±0.2 °2θ.The weak nature of the PXRD pattern indicates that Form III is poorlycrystalline. An image obtained by optical microscopy of Form III showedthe presence of some crystalline material with an irregular habit.

The Raman spectrum of Form III was obtained in a similar manner asdescribed in Example 1.2 for Form I. An expanded region of the Ramanspectrum of Form III (sample PP502-P120) is shown in FIG. 22 andexhibits peaks (Raman shift, cm⁻¹±2 cm⁻¹) at 1668, 1609, 1562, 1535,1494, 1450, 1350, 1324, 1306, 1264, 1245, 1190, 997, and 272.

The IR spectrum of Form III was obtained using the same method asdescribed in Example 1.2 for Form I. FIG. 23 shows the IR spectrum ofForm III (sample PP502-P120), and an expansion of the spectral regionfrom 1800 to 600 cm⁻¹ is shown in FIG. 24. The characteristic peaksobserved for Form III are provided in Table 7.

TABLE 7 Characteristic peaks for Form III determined by ATR IRspectroscopy (+/−4 cm⁻¹). Wavenumber Intensity (cm⁻¹) (arbitrary units)3446 0.159 2248 0.048 1667 0.292 1592 0.536 1531 0.352 1504 0.400 14280.659 1349 0.171 1305 0.507 1243 0.252 1189 0.160 1158 0.159 1089 0.1301001 0.235 896 0.181 862 0.197 829 0.164 780 0.362 759 0.280 736 0.424699 0.342

The DSC thermogram of Form III is shown in FIG. 25, with events at 147°C. (−23.7 J/g) and 215° C. (141 J/g) assigned to solvent loss andmelting, respectively. By TGA, Form III was observed to lose 4.8% massover the temperature range of 40-130° C.

Because it was known that Form III is a metastable hydrate, the DVSanalysis of Form III (sample PP502-P120) was programmed to begin withincreasing relative humidity instead of decreasing relative humidity.The experiment began at 50% RH, which was increased to 95% RH, decreasedto 0% RH, and finally increased back to the starting 50% RH. Theresulting DVS shows a maximum water content of about 8.5% at 95% RH andnearly all of the water is removed at 0% RH. The DVS results, includingsorption and desorption isotherm curves, show the total weight gainobserved between 0% RH and 80% RH to be about 8%, which indicates thatForm III is hygroscopic according to the EP classification (see Example1.2). Little hysteresis is observed between the sorption and desorptioncurves. The DVS results, combined with PXRD data taken before and afterthe DVS experiments, indicates that Form III is a non-stoichiometricchannel hydrate, rather than a dihydrate, because the water content canvary continuously over the whole relative humidity range. Like Form II,Form III thus also behaves as a variable hydrate.

Example 4 Forms Prepared From Form II (Forms IV-VIII of the Free Base ofFormula (1)

In addition to Form II, which is a trihydrate with a typical watercontent of about 10%, several other derivatives of Form II were alsoinvestigated. For example, when Form II is dehydrated below about 20%relative humidity (RH), another non-solvated form is obtained. This formis designated as Form IV. Characterization of Form IV was carried outusing various techniques including PXRD and DSC, which were performed asdescribed previously for the characterization of Forms I and II.

In order to evaluate the state of dehydrated Form II, a sample of thetrihydrate (Form II) was placed into a 1.0 mm PXRD sample holder andkept under dry nitrogen overnight. After 24 hours, the sample holder wascovered with a PMMA dome to keep the sample under nitrogen, and a PXRDpattern was recorded. FIG. 26 depicts the PXRD pattern for Form IV. Thefollowing peaks were identified in the PXRD pattern of FIG. 26: 7.0,8.5, 9.6, 10.3, 11.5, 11.9, 14.3, 14.9, 16.1, 17.0, 18.2, 19.3, 20.2,20.6, 21.1, 21.6, 22.1, 22.8, 23.1, 24.0, 25.4, 26.9, 27.6, 28.4, 28.7,29.3, 30.4, 31.8, 32.5, 33.5, 33.9, and 34.9 °2θ±0.2 °2θ.

Because the DVS results of Form II indicate reversible moisturesorption-desorption behavior (see above), tests were performed toconfirm that storage of the dehydrated Form II sample (i.e., Form IV) atabout 60% RH would again lead to the trihydrate form (Form II).Reexamination by PXRD confirms that dehydrated Form IV does revert toForm II after storage at 60% RH for three days.

DSC analysis of Form IV was performed after the sample was equilibratedunder dry nitrogen for about 60 hours. The dehydrated sample was exposedto temperature ramp rates of 10° C./minute or 20° C./minute up toapproximately 240° C. The DSC thermogram illustrates a melting peak atapproximately 159° C. with an enthalpy of fusion of about 57 J/g.Thermal decomposition begins immediately after melting.

Another dehydrated form was obtained when Form II was dried at 100° C.under vacuum for 2 hours. This form is designated as Form V. From DVSanalysis of Form II (see above), it is known that Form II loses waterwhen kept under dry nitrogen. Form V was identified while studying thebehavior of Form II after exposure to elevated temperatures.Characterization of Form V was carried out using various techniquesincluding: PXRD (FIG. 27); and Raman spectroscopy (FIG. 28). Form Vshows a distinct PXRD pattern and a new Raman spectrum, which wereperformed as described previously for the characterization of Forms Iand II. ¹H NMR spectroscopy confirmed the chemical integrity of thecompound.

FIG. 27 shows the PXRD pattern for Form V, and specifically, samplePP502-P44. The following peaks were identified in the PXRD pattern ofFIG. 27: 4.5, 5.5, 5.9, 8.1, 10.6, 11.1, 11.9, 13.2, 17.9, 19.2, 19.9,20.4, 21.3, 21.8, 22.6, 23.7, 24.3, 24.7, 25.0, 26.0, 26.3, 27.6, 28.6,and 30 °2θ±0.2 °2θ. Comparisons between the PXRD pattern for Form II(FIG. 12) and the PXRD pattern for Form V show overlapping peaks at11.0, 19.3, 22.0, 22.5, 22.7, 23.9, 24.2, 24.6, 25.0, 26.1, 27.5 °2θ±0.2°2θ for Form II and 11.1, 19.2, 21.8, 22.6, 23.9, 24.3, 24.7, 25.0,26.0, 27.6 °2θ±0.2 °2θ for Form V with the following peaks of Form IIeither disappearing entirely or decreasing in intensity: 9.9, 11.0,14.3, 18.1, 18.4, 18.9, 20.2, 22.0, 22.2, 22.5, 22.7, 22.9, 23.9, 24.6,26.1, 26.6, 28.2, and 32.7 °2θ.

Thermoanalytical characterization of Form V was carried out usingTG-FTIR and DSC analytical techniques. The TG-FTIR thermogram shows thatthe sample immediately loses about 5% of its water mass upon heating ata rate of 10° C. per minute to about 100° C. to 120° C. The sampleremains stable till approximately 200° C., at which point an additionalmass change of about 17% is seen due to sample decomposition uponcontinued heating up to approximately 340° C. to 350° C.

DSC of a Form V sample was carried out in a sample pan sealed underambient conditions. The DSC thermogram shows a very broad endotherm witha peak at about 125° C. A substantial part of this endothermic signalcorresponds to release of water from the sample into the void volume ofthe hermetically sealed sample pan since the TG-FTIR thermogram showsthat the release of water commences just above ambient temperature,indicating that the water is likely to be loosely bound in the crystalstructure.

The FT-Raman spectrum of Form V in the relevant fingerprint region (200cm⁻¹ to 1800 cm⁻¹) is shown in FIG. 28 and exhibits peaks (Raman shift,cm⁻¹±2 cm⁻¹) at 1686, 1613, 1574, 1540, 1504, 1488, 1349, 1314, 1288,1266, 1193, 1153, 1052, 1027, 852, 775, 708, and 378. Differences in theRaman spectra of Form V and Form II are highlighted by peaks at 1686,1574, 1488, 1314, 1266, 1193, 1153, 1052, and 708 cm⁻¹ of the Form Vspectra, all of which do not appear in the Form II spectrum (see FIG.15). Furthermore, peaks appearing at 1668, 1580, 1564, 1493, 1454, 1436,1416, 1401, 1321, 1272, 1252, 1244, 1183, 1165, 1097, 1039, 996, 950,871, 730, 645, 633, 352, 279, and 247 cm⁻¹ of the Form II spectrum arenot present in the Form V spectra, indicating the presence of distinctphases.

A comparison of the Raman spectra of Forms I, II and V, as shown inFIGS. 4, 14 and 28, respectively, illustrates that all three forms maybe readily distinguished by Raman spectroscopy.

Crystallization of Formula (1) in methanol and methanol-water mixtures(95:5) led to samples with new PXRD patterns (samples P502-P26 andPP502-P16, respectively). Form VI was the product from a crystallizationexperiment conducted in a methanol-water mixture (95:5) at 5° C. (samplePP502-P16). Form VI exhibits a unique PXRD pattern (FIG. 29) and Ramanspectrum (FIG. 30). Comparison of the PXRD pattern for Form VI (FIG. 29)with those of Form I (FIG. 1) and Form II (FIG. 12) shows that neitherForm I nor Form II is present in sample PP502-P16. The followingcharacteristic peaks were identified in the PXRD pattern of Form VI:6.5, 6.8, 8.5, 11.8, 12.6, 13.5, 13.8, 14.8, 15.0, 16.2, 16.4, 16.9,18.5, 19.4, 19.9, 20.6, 21.6, 22.1, 22.7, 23.6, 24.5, 24.8, 25.3, 26.0,27.2, 27.8, 28.5, 28.9, 30.2, and 34.3 °2θ±0.2°2θ. Characteristic peaksfor Form VI in FIG. 31 are observed at 1667, 1609, 1580, 1562, 1535,1495, 1450, 1350, 1323, 1306, 1264, 1245, 1190, 1161, 1042, 997, 838,762, 717, 630, and 272 cm⁻¹±2 cm⁻¹.

Form VII was obtained from the crystallization experiment conducted inpure methanol where the obtained solid sample PP502-P26 precipitated outwhen stored at 4° C. Form VII exhibits a unique PXRD pattern (FIG. 31)and Raman spectrum (FIG. 32). Comparison of the PXRD pattern for FormVII with those of Form I and Form II (FIG. 31, FIG. 1 and FIG. 12,respectively) shows that neither Form I nor Form II is present in samplePP502-P26. The following characteristic peaks were identified in thePXRD pattern of Form VII: 5.9, 6.5, 7.0, 7.8, 8.5, 9.6, 10.0, 10.4,13.4, 13.9, 15.0, 16.5, 16.9, 17.7, 18.5, 19.0, 19.9, 20.8, 21.6, 22.4,23.7, 23.9, 24.8, 25.2, 27.5, 28.3, and 30.0 °2θ±0.2 °2θ. CharacteristicRaman peaks for Form VII in FIG. 32 are observed at 1687, 1663, 1604,1578, 1561, 1534, 1486, 1462, 1443, 1397, 1361, 1348, 1327, 1305, 1251,1234, 1184, 1163, 1037, 1001, 835, 774, 757, 717, 653, 606, 422, 348,and 268 cm⁻¹±2 cm⁻¹.

¹H NMR spectroscopy shows that both Form VI and VII contain about 0.7equivalents of methanol, and according to TG-FTIR, both forms alsocontain substantial amounts of water. Forms VI and VII are likely to bemetastable methanol solvates or mixed solvate-hydrates (i.e.,methanolate-hydrate). Suspension equilibration experiments conducted at5° C. in methanol with a mixture of Forms (I), VI, and VII show thatpure Form I was recovered after five days, suggesting that even in puremethanol, Form I is likely to be more stable than either Form VI or FormVII.

Form VIII (sample PP502-P23) is a putative acetic acid disolvate. ThePXRD pattern of Form VIII is shown in FIG. 33. The following peaks wereidentified in the PXRD pattern of Form VIII: 4.28, 6.18, 6.59, 8.57,11.75, 12.03, 12.38, 15.61, 17.16, 18.02, 18.58, 19.43, 20.02, 20.85,22.07, 22.68, 23.69, 25.13, 25.96, 26.43, and 27.49 °2θ±0.2 °2θ. TheRaman spectrum of Form VIII is shown in FIG. 34. Characteristic Ramanpeaks for Form VIII are observed at 1681, 1580, 1529, 1497, 1456, 1437,1349, 1313, 1302, 1268, 1243, 1193, 1157, 1047, 1025, 1006, 951, 896,851, 775, and 264 cm⁻¹±2 cm⁻¹.

An overview of the forms is presented in Table 8.

TABLE 8 Overview of crystalline forms of the free base of Formula (1).Stability at Room Form Description Temperature Comments Form Ianhydrous, stable relative to Stability at lower a_(w) non-solvated FormII below enables isolation of Form I a_(w) = 0.6 from organic solventsForm II trihydrate stable relative to Variable hydrate; contains Form Iabove up to about 10% water at a_(w) = 0.6 variable levels Form IIIdihydrate metastable Higher energy variable hydrate; contains up toabout 8% water at variable levels Form IV anhydrous, metastable Form IIdehydrated by low non-solvated moisture Form V anhydrous, metastableForm II dehydrated by heat non-solvated Form VI methanol metastableOrganic solvate solvate Form VII methanol metastable Organic solvatesolvate Form VIII acetic acid metastable Organic solvate disolvate

A diagram showing the transformation scheme between the forms is givenin FIG. 35. In FIG. 35, black arrows indicate that the transformationhas been established by experiments, while blue arrows indicate that thetransformation may occur.

Example 5 Preparation and Characterization of Amorphous(S)-4-(8-Amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide

Amorphous(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidecan be prepared by various methods, including the procedure described inExample 6 of U.S. Patent Application Publication No US 2014/0155385 A1and International Patent Application Publication No. WO 2013/010868 A1,the disclosure of which is incorporated herein by reference. Fastevaporation of the solvent from a solution in dichloromethane or in amixture of dichloromethane with a co-solvent, e.g., acetone or analcohol, can be used to prepare the amorphous form. In addition, theamorphous form can be produced by freeze drying of an aqueous solutionthat contains a small amount of an acid, e.g., formic acid or aceticacid, to solubilize the free base form I.

Amorphous(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidecan be prepared by adding 3.0 mL of water to 200 mg of crystalline FormI. Formic acid is then added dropwise until dissolution of the solid iscomplete. About 50 microliter of formic acid is typically sufficient toachieve complete dissolution of the form I. The clear solution isfiltered into a 100 mL round flask through a 0.22 μm microporepolytetrafluoroethylene (PTFE) filter (for instance using a syringe) andthe solution in the round flask is freeze dried. The resulting product(sample PP502-P107) is the amorphous form. Characterization of theproduct after freeze drying by powder X-ray diffraction reveals that theamorphous form is obtained. The resulting PXRD pattern is shown in FIG.36. No Bragg reflections are observed, and the PXRD pattern ischaracterized by diffuse scattering typical of an amorphous material.

An expanded region of the Raman spectrum of a similarly-prepared sampleof amorphous Formula (1) (sample PP502-P118) is shown in FIG. 37.Characteristic Raman peaks are observed at 1674, 1608, 1577, 1537, 1492,1449, 1348, 1307, 1238, 1188, and 992+/−4 cm⁻¹.

An expanded region of the IR spectrum of a similarly-prepared sample ofamorphous Formula (1) (sample P502-P148) obtained with ATR sampling isshown in FIG. 38. The y-axis is shown in arbitrary units with atransmittance scale. Characteristic IR peaks are observed at 1668, 1605,1505, 1428, 1302, 1237, 1200, 1153, 1091, 997, 944, 894, 863, 776, and735+/−4 cm⁻¹ which are distinct from the spectra of other crystallineforms of Formula (1).

Further characterization of the product after freeze drying by TG-FTIRrevealed that a small amount of formic acid is present. Therefore, theobtained amorphous sample was further dried under vacuum at 80° C. forabout 20 hours and retested by TG-FTIR and DSC. The TG-FTIR and DSCthermograms of sample PP502-P107A are shown in FIG. 39 and FIG. 40,respectively. TG-FTIR of the dried sample showed that only very littlewater and residual solvent was present. DSC of the essentiallysolvent-free amorphous form shows a glass transition temperature atabout 130° C. with a ΔC_(p) of about 0.3 J/(g·K). A smaller change ofthe heat capacity at about 120° C. might be due to another fraction ofamorphous material that might contain traces of solvents, thus showing areduced glass transition temperature. Thermal degradation begins aboveabout 160° C.

The DVS results for a similarly-prepared sample of amorphous Formula (1)(sample P502-P148), including sorption and desorption curves, wasperformed by exposing samples to a starting 50% RH, decreasing humidityto 0% RH, increasing humidity to 95% RH, and finally decreasing humidityback to the starting 50% RH. The DVS results, including sorption anddesorption curves, show that significant water gain occurs duringsorption starting at about 20% RH. The total water gain observed between0% RH and 80% RH is about 6% by weight, which indicates that Form II ishygroscopic according to the EP classification (see Example 1.2).Furthermore, the total mass gained up to 95% RH is approximately 13%,and the mass is irreveribily gained after sorption (with the finaldecrease to 50% RH only removing 5% of the 10% moisture gained in the 50to 95% RH range). The results illustrate the hygroscopic nature of theamorphous form, including the irreversible uptake of a large amount ofwater upon exposure to high RH.

Example 6 Crystalline Salts of(S)-4-(8-Amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide

A salt screening with(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base (sample PP502-P1, prepared as described above, comprisedcrystallization experiments with 11 different acids, including: benzoicacid, benzenesulfonic acid, citric acid, fumaric acid, hydrochloricacid, maleic acid, nicotinic acid, phosphoric acid, saccharin, succinicacid, and L-tartaric acid. Of these, crystalline samples were obtainedwith citric acid, fumaric acid, maleic acid, phosphoric acid, succinicacid and L-tartaric acid.

A summary of the starting materials for the salt preparations isprovided in Table 9. Each prepared product was given a sample identifieras follows: SP221-XXX-Pn (XXX=salt identification code, andn=experiment/sample number).

TABLE 9 Summary of starting materials for salt preparation. m SampleCompound pK_(a) [g/mol] Source/Number Designation Free base ~5.7 465.5Formula (1)/ PP502-P1 CML1476, Lot CS13-083 HB873-98 Fumaric 3.0 (4.4)116.07 Sigma # 240745 SP221-FUM-Pn acid Maleic acid 1.9 (6.2) 116.1Fluka # 63180 SP221-MLE-Pn Phosphoric 2.0 (7.1) 98.0 Fluka # 79606SP221-PO4-Pn acid L-Tartaric 3.0 (4.4) 150.09 Fluka # 95310 SP221-LTA-Pnacid

Characterization of fumarate salt, maleate salt, phosphate salt andL-tartrate salt was conducted using ¹H NMR spectroscopy, TG-FTIR, DSC,dynamic vapor sorption, optical microscopy, high-performance liquidchromatography (HPLC) purity, laser diffraction, approximate bulk andtapped density, and aqueous solubility analytical techniques.

Once seeding crystals were obtained, the formation of crystalline saltswas shown to be reproducible, with the various salts showing a goodtendency towards crystallization. A summary of salt properties isdetailed in Table 10.

TABLE 10 Summary of Formula (1) salt properties in comparison to freebase. Melting Pt./ Solubility Thermal Stability Solid Form Salt (3hours) (Decomposition Pt.) Behavior in DVS Assessment Free base S < 1mg/mL 215° C., (Form I) Form I is not Anhydrate Form I hygroscopic;higher and trihydrate melting point Form II Fumarate ~1.8 mg/mL ~170°C./170° C. Reversible anhydrate- Sesquihydrate, hydrate formation,possibly multiple Δm~0.8% (20-80% RH) forms Maleate 2.2 mg/mL ~161°C./170° C. Water more strongly Possibly sesqui- bound than in thehydrate, multiple fumarate, Δm~0.7% forms (20-80% RH) Phosphate 9.8mg/mL ~157° C./180° C. Reversible anhydrate- At least two forms, hydrateformation, anhydrate and Δm~0.7% (20-80% RH) hydrate L-Tartrate 5.3mg/mL ~158° C./165° C. Does not completely Possibly sesqui- dehydrate at0% RH, hydrate, possibly Δm~0.7% (20-80% RH) multiple forms

Because the solubility was measured after an equilibration time of onlythree hours and without adjustment of pH, the solubility of all salts isdramatically increased in comparison to the free base. Whereas the freebase is a poor water soluble drug, the salts were well soluble in water.

All of the salts described above appear to form hydrates. Preliminarytest experiments consisted of one to three suspension equilibrationexperiments for each salt. Results show that all four salts can exist inmultiple solid forms including polymorphic forms.

Example 6.1 Form A of the Fumarate Salt of Formula (1)

Crystalline Form A of the fumarate salt of Formula (1) was prepared bydissolving 16.294 g of PP502-P1 free base and 4.065 g fumaric acid in500 mL acetone. The mixture was subsequently heated to 50° C., whereby50 mL of water was added. The water addition led to a clear solution atwhich time, the solution was allowed to cool to room temperature whilestirring at about 300 rpm. At room temperature, the clear solution wasseeded with about 20 mg of SP221-FUM-P5 and after about 48 hours, thesuspension was filtered to obtain a solid which was dried in air at 40°C. for about 24 hours. Initial characterization of the obtained solidresulted in a yield of approximately 13.6 grams (about 64%) at about99.9% purity, as measured by high-performance liquid chromatography(HPLC).

Form A of the fumarate salt was characterized by ¹H NMR spectroscopy;optical microscopy, Fraunhofer laser diffraction, reflection PXRD (FIG.41), TG-FTIR, DSC and dynamic vapor sorption (DVS).

The ¹H NMR spectrum of the fumarate salt (sample SP221-FUM-P9) wasrecorded in acetone solvent and confirmed the composition is consistentwith a 1:1 fumarate salt. A peak near 2.1 ppm indicates a trace ofacetone as residual solvent that is still present after drying.

The fumarate salt was obtained as small particles. Because the obtainedsalt was clumpy, the dried material was sieved through a 500 μm sieveprior to further characterization. Examination by polarized lightoptical microscopy was conducted by dispersing the compound in heptaneand thereafter sonicating for short period of time sufficient todisperse the crystals. Optical microscopy revealed very smallcrystalline particles that, after sieving and dispersion, are stilllargely agglomerated.

Particle size distribution testing was conducted using Fraunhofer laserdiffraction with values for the maximum particle size for a givenpercentage volume of the sample shown in Table 11 below. For example,the size dimension at ×50 (42 μm) represents the maximum particlediameter below which 50% of the sample volume exists. This parameter isalso known as the median particle size by volume.

TABLE 11 Particle size distribution results for the fumarate salt.Sample x10 x50 (median) x90 SP221-FUM-P9a 3.6 μm 42 μm 329 μm

By monitoring these three parameters (×10, ×50, ×90), it is possible todetermine if there are significant changes in the main particle size, aswell as changes at the extremes of the distribution, possibly due to thepresence of fines or oversized particles or agglomerates in the particlesize distribution. Static light scattering techniques such as laserdiffraction give a volume weighted distribution wherein the contributionof each particle in the distribution relates to the volume of thatparticle (equivalent to mass if the density is uniform). This isextremely useful since the distribution represents the composition ofthe sample in terms of its volume/mass.

PXRD, along with the optical microscopy images, confirmed thecrystalline nature of the salt. The relection PXRD pattern of fumaratesalt sample SP221-FUM-P9 is depicted in FIG. 41 and shows the followingrepresentative peaks at: 4.9, 5.4, 7.0, 9.8, 10.8, 11.5, 12.1, 14.1,16.1, 16.6, 17.8, 18.5, 19.4, 20.3, 2056, 21.9, 22.1, 22.5, 23.1, 24.0,24.8, 26.6, 26.8, 27.3, and 28.2±0.2 °2θ.

Thermoanalytical characterization of Form A of the fumarate salt wascarried out using TG-FTIR and DSC. TG-FTIR analysis of a representativecrystalline Formula (1) fumarate sample revealed a mass loss of about4.5%; this is essentially attributable to water loss. The amount ofwater attributed to the loss in mass closely matches the theoreticalwater content of a sesquihydrate at 4.6%. Mass loss of about 12.75% uponheating thereafter to approximately 300° C. was due primarily todecomposition. DSC of the same sample shows an endothermic peak near162° C. that deviates from the baseline above about 120° C. andincreases slowly. However, because of an exothermic degradationbeginning at around 170° C., the enthalpy of fusion cannot be evaluatedreliably.

Hygroscopic behavior of the fumarate salt (sample SP221-FUM-P9a) wasmeasured using dynamic vapor sorption. The DVS sorption and desorptionresults indicate that the salt loses nearly all water content at lowhumidity conditions while reaching a maximum saturation of approximately6% at a RH of about 95%. The water content change between 20% and 80% RHis about 0.8%. Similar to previously described moistureadsorption-desorption isotherms, samples were exposed to a starting 50%RH, decreasing humidity to 0% RH, increasing humidity to 95% RH, andfinally decreasing humidity back to the starting 50% RH.

Example 6.2 Form A of the Maleate Salt of Formula (1)

Crystalline Form A maleate salt (sample SP221-MLE-P9) was prepared bydissolving 16.296 g of PP502-P1 free base in a 350 mL of acetone and 35mL of water mixture. The mixture was subsequently heated to 50° C.,which led to a clear solution. Thereafter, 20 mL of an aqueous solutioncontaining 4.043 g of maleic acid was added. Moreover, the vessel andpipette holding the aqueous maleic acid solution were washed with 1.0 mLof water and the wash solution was also included in the mixture. Thesolution was allowed to cool while stirring at about 300 rpm. At about45° C., the solution was seeded with about 20 mg of SP221-MLE-8 andfurther cooled to approximately 20° C. After about 24 hours, thesuspension was filtered to obtain a solid which was dried in air at 40°C. for about 20 hours. Initial characterization of the obtained solidresulted in a yield of approximately 14.1 grams (about 66%).

The maleate salt was characterized by ¹H NMR spectroscopy, opticalmicroscopy, Fraunhofer laser diffraction, reflection PXRD (FIG. 42),TG-FTIR, DSC, and DVS.

The ¹H NMR spectrum of the maleate salt (sample SP221-MLE-P9), recordedin acetone solvent, confirmed the composition and is consistent with a1:1 maleate salt. Similar to the ¹H NMR analysis of the fumarate salt ofFormula (1), a minute trace (0.7%) of acetone as residual solvent isalso present in the spectra.

Examination by polarized optical microscopy revealed that the maleatesalt consists of small crystalline particles ranging in size from about10 μm to about 100 μm. The prepared maleate salt was substantiallylarger than the particles of the fumarate salt (discussed above), thephosphate salt and the L-tartrate salt (latter two discussed below). Thefine powder showed favorable flow properties and no sieving wasnecessary after drying.

Particle size distribution testing was conducted using Fraunhofer laserdiffraction with values for the maximum particle size for a givenpercentage volume of the sample shown in Table 12 below.

TABLE 12 Particle size distribution results for the maleate salt. Samplex10 x50 (median) x90 SP221-MLE-P9 10.7 μm 38 μm 73 μm

Particle size distribution function for the maleate salt and opticalmicroscopy images confirm a particle size distribution ranging roughlybetween 10 μm and 100 μm.

Powder X-ray diffraction, along with the optical microscopy, confirmedthe crystalline nature of the salt. The reflection PXRD pattern ofmaleate salt sample SP221-MLE-P9 is depicted in FIG. 42 and shows thefollowing representative peaks at: 5.3, 9.8, 10.6, 11.6, 13.5, 13.8,13.9, 14.3, 15.3, 15.6, 15.8, 15.9, 16.6, 17.4, 17.5, 18.7, 19.3, 19.6,19.8, 20.0, 20.9, 21.3, 22.0, 22.3, 22.7, 23.2, 23.4, 23.7, 23.9, 24.5,24.8, 25.2, 25.6, 26.1, 26.4, 26.7, 26.9, 27.1, 27.6, 28.8, 29.5, 30.0,30.3, 30.9, 31.5, 31.9, 32.5, 34.0, and 35.1 °2θ±0.2 °2θ.

Thermoanalytical characterization of the maleate salt was carried outusing TG-FTIR, and DSC. TG-FTIR analysis of the representativecrystalline Formula (1) maleate sample revealed a mass loss of about5.3%; this is essentially attributable to water loss. The amount ofwater attributed to the loss in mass closely matches the theoreticalwater content of sesquihydrates at 4.6%. However, no acetone wasdetected. Mass loss of about 10.1% upon heating thereafter toapproximately 300° C. was due primarily to decomposition. DSC of thesame sample shows an endothermic peak near 174° C., followed bydecomposition.

Hygroscopic behavior of the maleate salt (sample SP221-MLE-P9) wasmeasured using dynamic vapor sorption. The DVS sorption and desorptionresults indicate that the salt loses very little water at 0% RH. Thesample reaches a maximum saturation of approximately 5.8% at a RH ofabout 95%. The water content change between 20% and 80% RH is about0.5%. Samples were exposed to a starting 50% RH, decreasing humidity to0% RH, increasing humidity to 95% RH, and finally decreasing humidityback to the starting 50% RH.

Example 6.3 Form A of the Phosphate Salt of Formula (1)

Preparation of Form A of the phosphate salt of Formula (1) wasaccomplished as follows. First, 350 mL of acetone and 35 mL of waterwere added to 16.2998 grams (35 mmol) of Formula (1) free base PP502-P1.Upon heating to 50° C., a clear solution was obtained. To this solutionwas slowly added 2.5 mL of 85-90% phosphoric acid (35 mmol). Thesolution was allowed to cool while stirring at about 300 rpm. At about38° C., crystallization was observed without seeding. After about 80hours the suspension was filtered and the obtained solid was dried inair at 40° C. for about 24 hours. The yield was approximately 20.48grams (97%). The phosphate salt was obtained as small particles. Afterdrying the material was clumpy and very strongly agglomerated particleswere observed. In order to obtain a free flowing powder for the tappeddensity test and particle size analysis, the dried material was sievedthrough a 500 μm sieve. The sample after drying was termed SP221-PO4-P5and the sample after sieving was termed SP221-PO4-P5a.

Initial characterization of the obtained solid resulted in a purity ofabout 99.9%, as measured by HPLC. Based on DVS and TG-FTIR, thephosphate salt form produced is likely to be a dihydrate having atheoretical phosphorus content of about 5.2%. The phosphorus content wasexamined by inductively-coupled plasma optical emission spectrometry(ICP-OES) and was determined to be approximately 4.7%, which is slightlybelow the required content for a 1:1 salt.

The phosphate salt was characterized by ¹H NMR spectroscopy, opticalmicroscopy, Fraunhofer laser diffraction, reflection PXRD (FIG. 43),TG-FTIR, DSC, and DVS.

¹H NMR spectroscopy confirmed the composition was consistent with thestructure of a crystalline phosphate salt. Examination by polarizedoptical microscopy revealed that the phosphate salt was a crystallinematerial that consisted of very small particles, most of which were lessthan about 10 μm in diameter. Sample SP221-PO4-P4 shows needle-shapedparticles. While needles are not clearly visible for sampleSP221-PO4-P5, one reason might be because of the small size of theparticles.

Particle size distribution testing was conducted using Fraunhofer laserdiffraction with values for the maximum particle size for a givenpercentage volume of the sample shown in Table 13 below.

TABLE 13 Particle size distribution results for the phosphate salt.Sample x10 x50 (median) x90 SP221-PO4-P5 2.8 μm 29 μm 191 μm

PXRD confirmed the crystalline nature of the salt. The reflection PXRDpattern of a sample taken from a 20 gram batch of sample SP221-PO4-P5phosphate salt is depicted in FIG. 43 and shows the followingrepresentative peaks at: 4.5, 6.0, 7.2, 10.4, 12.0, 12.5, 13.1, 14.3,15.5, 17.4, 18.0, 18.3, 18.9, 19.3, 20.2, 20.5, 20.9, 21.4, 21.9, 22.0,22.6, 22.9, 23.1, 23.3, 24.2, 24.6, 25.0, 25.7, 26.2, 26.4, 26.9, 27.3,27.5, 29.3, 30.0, 30.3, 30.5, 30.9, 31.2, 31.9, and 35.7 °2θ±0.2 °2θ.The phosphate salt exists in at least two different crystalline forms,an anhydrous form of the crystalline structure and a hydrate form of thecrystalline structure, with each form exhibiting unique PXRD patterns.The peaks of FIG. 44 correspond to the hydrate form of the crystallinephosphate salt.

Thermoanalytical characterization of the phosphate salt was carried outusing TG-FTIR and DSC. TG-FTIR analysis of the crystalline Formula (1)phosphate salt (sample SP221-PO4-P1) revealed a mass loss of about 5.9%;this is essentially attributable to water loss. This result suggeststhat the obtained crystalline form of the phosphate is a dihydrate sincethe 5.9% water content of the phosphate sample is close to the expectedcontent for a dihydrate (6.0%). Additional mass losses upon heatingthereafter to approximately 250° C. were due primarily to decomposition.DSC of the same sample shows a broad endothermic peak near 138° C. Theenthalpy of fusion is estimated to about 134 J/g.

Hygroscopic behavior of the phosphate salt (sample SP221-PO4-P1) wasmeasured using DVS. The DVS sorption and desorption results indicatethat the salt loses nearly all water content at low RH conditions whilereaching a reaching a maximum saturation of approximately 6.6% at a RHof about 95%. DVS analysis suggests that the phosphate salt forms adihydrate with a water content of about 6.0%. Samples were exposed to astarting 50% RH, decreasing humidity to 0% RH, increasing humidity to95% RH, and finally decreasing humidity back to the starting 50% RH.

Example 6.4 Form A of the L-Tartrate Salt of Formula (1)

Crystalline Form A of Formula (1) L-tartrate salt was prepared bydissolving 16.298 g of PP502-P1 free base in a 350 mL of acetone and 35mL of water mixture. The mixture was subsequently heated to 50° C.,which led to a clear solution. Thereafter, 20 mL of an aqueous solutioncontaining 5.257 g of L-tartaric acid was added to the clear solution.The solution was allowed to cool to approximately 20° C. while stirringat about 300 rpm. After about 24 hours, the suspension was filtered toobtain a solid which was dried in air at 40° C. for about 20 hours.Initial characterization of the obtained solid resulted in a yield ofapproximately 20.1 grams (about 89%) at about a 99.78% purity, asmeasured by HPLC.

The L-tartrate salt was characterized by ¹H NMR spectroscopy, opticalmicroscopy, Fraunhofer laser diffraction, reflection PXRD (FIG. 44),TG-FTIR, DSC, and DVS.

¹H NMR spectroscopy confirmed the composition was consistent with thestructure of a 1:1 crystalline L-tartrate salt. The L-tartrate salt wasobtained as a crystalline material; examination by polarized opticalmicroscopy revealed that the material consists of partially agglomeratedfine needles varying in length from about 2 to about 40 μm and widths onthe order of a few μm.

Particle size distribution testing was conducted using Fraunhofer laserdiffraction with values for the maximum particle size for a givenpercentage volume of the sample shown in Table 14 below.

TABLE 14 Particle size distribution results for the L-tartrate salt.Sample x10 x50 (median) x90 SP221-LTA-P8a 1.7 μm 17 μm 59 μm

PXRD, along with the optical microscopy, confirmed the crystallinenature of the salt. The reflection PXRD pattern of L-tartrate saltsample SP221-LTA-P8 is depicted in FIG. 80 and shows the followingrepresentative peaks at: 4.6, 5.5, 7.2, 9.3, 10.7, 10.9, 11.8, 14.3,14.9, 16.4, 17.0, 17.7, 19.2, 19.4, 19.5, 20.3, 21.6, 22.4, 23.3, 23.8,24.3, 24.5, 24.7, 25.1, 25.6, 26.8, 27.2, 27.8, 28.4, 28.7, 29.0, 29.5,30.0, 30.9, 31.6, 32.1, 32.4, 33.0, 33.5, and 33.9 °2θ±0.2 °2θ.

Thermoanalytical characterization of the L-tartrate salt was carried outusing TG-FTIR and DSC. TG-FTIR analysis of the crystalline Formula (1)L-tartrate salt (sample SP221-LTA-P8) revealed a mass loss of about4.8%; this is essentially attributable to water loss. This amount ofwater is near the theoretical amount of water for a sesquihydrate whichis 4.3%. Additional mass loss of about 20% upon heating thereafter toapproximately 300° C. was due primarily to decomposition. DSC of thesame sample shows an endothermic peak near 156.5° C. with an enthalpy offusion of about 40.70 J/g.

Hygroscopic behavior of the L-tartrate salt (sample SP221-LTA-P8a) wasmeasured using dynamic vapor sorption. The DVS sorption and desorptionresults indicate that the salt loses water at low humidity conditionswhile reaching a maximum saturation of approximately 5.4% at a RH ofabout 95%. Moreover, from an initial water content of about 4.8% at 50%RH (confirmed by TG-FTIR), the DVS analysis suggests that about 30% ofthis water was removed within the timescale of the measurement. Thewater content change between 20% and 80% RH is about 0.7%. Moistureadsorption-desorption isotherms were prepared in a similar manner asdescribed above.

Example 7 Solubility as a Function of pH Example 7.1 Free BaseSolubility

The aqueous solubility of Formula (1) free base was examined as afunction of pH. Experiments were conducted in aqueous HCl solution andbuffer solutions at pHs of 1, 3, 5, 6.8, 7.4 and 9. It was determinedthat at low pH values of 1 and 3, the solid completely dissolved overthe equilibration time while the pH in the system stabilized toapproximately 3 in both experiments. It was found that the solubility inHCl solution having a pH of about 1 is at least 150 mg/mL. Solubility ofthe free base Form I (PP502-P1) at various pH values greater than 3 ispresented in Table 15.

TABLE 15 Solubility data for Formula (1) free base. Effective pH at theend of the test Solubility (mg/mL) 5.0 0.69 6.7 0.056 7.3 0.049 8.90.051

Data shows that aqueous solubility of Formula (1) free base (samplePP502-P1) stabilizes at approximately 50 μg/mL at about pH 6.7. FIG. 45displays the possible species of Formula (1) based on the calculatedpK_(a) values ranging from 2.2 (basic) to 6.1 (basic) to 11.5 (acidic).Therefore, a doubly-positive charged molecule is highly soluble inwater, whereas the singly positive and the neutral forms are poorlysoluble. This highlights the challenges in successful delivery ofFormula (1) through the stomach into the higher pH environment of theduodenum.

The equilibrium pH-solubility relationship calculated for the free baseof Formula (1) is shown in FIG. 46 in comparison to experimentalmeasurements at selected intervals. At pH 6.7 and greater, the aqueoussolubility reaches a constant level of about 50 μg/mL, furtherillustrates the challenges of delivery of Formula (1).

The pK_(a) values for Formula (1) free base were determined and used tocreate the speciation plot shown in FIG. 47 to illustrate the speciespresent as the Formula (1) passes through the gastrointestinal tract.The sample pK_(a) values were determined using an ultraviolet (UV)spectrometric technique. The sample was initially titrated in a fast-UVtriple titration between pH 2-12 at concentrations of 31-19 μM, underaqueous conditions. Three pK_(a) values, with average values of ˜3.6,˜5.8 and ˜12.0, were determined. The sample was subsequently titrated insix titrations, under aqueous conditions over a total range of pH1.5-12.5 at concentrations of 30-18 μM. Three pK_(a) values for Formula(1), with average values of 3.54±0.01, 5.77±0.01 and 12.12±0.03, weredetermined from the spectroscopic data collected.

The log P of Formula (1) free base was determined using thepotentiometric (pH-metric) technique. The sample was titrated in variousratios of octanol/water from pH 1.9-12.1 at concentrations of 1.1-0.5 mMat 25° C. in an ionic environment of 0.15 M KCl. The potentiometric datacollected were used to calculate the log P of the neutral (2.03±0.01)and the cationic (−0.31±0.06) species.

Example 7.2 Salt Solubility

Tests were directed to determine the aqueous solubility of the maleatesalt as a function of pH. Experiments were conducted in aqueous HClsolution and buffer solutions at pHs of 1, 3, 5, 6.8, 7.4 and 9. It wasdetermined that at low pH values of 1 and 3, the solid completelydissolved over the equilibration time while the pH in the systemstabilized to approximately 3 in both experiments. In parallel, aqueoussolubilities of the fumarate, maleate, phosphate, and L-tartrate wasdetermined in pure water. Solubility data for the fumarate, maleate,phosphate, and L-tartrate salts in pure water are presented in Table 16,with the phosphate salt showing the highest apparent solubility after 24hours. The solubility data for the maleate salt as a function of pH invarious aqueous media is presented in Table 17.

TABLE 16 Solubility data for selected salts in pure water at 25° C.Solubility (mg/mL) after 24 hours Salt Sample name pH equilibrationFumarate SP221-FUM-P9a 4.0 2.1 Maleate SP221-MLE-P9 4.1 2.3 PhosphateSP221-PO4-P5 3.6 15.0 L-tartrate SP221-LTA-P8 3.8 3.7

TABLE 17 Solubility data for the maleate salt as a function of pH inaqueous media. Solubility Sample name Description pH after 24 hours(mg/mL) SP221-MLE-P8_pH1 0.1N HCl, Sigma # 71763 1.8 37.0SP221-MLE-P8_pH3 citrate buffer pH 3, VWR #109434 3.2 8.0SP221-MLE-P8_pH5 buffer pH 5, Sigma # 33544 4.8 4.3 SP221-MLE-P8_pH6.8phosphate buffer pH 7, KH2PO4 4.8 1.2 SP221-MLE-P8_pH7.4 phosphatebuffer pH 7, KH2PO4 5.0 0.8 SP221-MLE-P8_pH9 borate buffer, VWR # 1094084.7 1.2

Example 7.3 Bulk and Tapped Density of Salts

The bulk and tapped density of the fumarate, maleate, phosphate, andL-tartrate salts were estimated by conducting experiments on four 10gram samples. The results are presented in Table 18.

TABLE 18 Bulk and tapped density measurements for selected salts. Tappeddensity Salt Sample name Bulk density (g/mL) (g/mL) FumarateSP221-FUM-P9a 0.45 0.57 Maleate SP221-MLE-P9 0.35 0.45 PhosphateSP221-PO4-P5a 0.33 0.48 L-Tartrate SP221-LTA-P8a 0.29 0.50

Example 8 Analytical Characterization of Form I Recrystallized fromEthanol (Sample P502-P99)

About 60 g of Formula (1) batch CS13-083 HB873-98 was recrystallizedfrom ethanol. The product was characterized by PXRD (FIG. 49), DSC,TG-FTIR, ¹H NMR spectroscopy, high-performance liquid chromatographypurity analysis, Fraunhofer laser diffraction, DVS, and opticalmicroscopy.

PXRD analysis confirmed that the structure of sample PP502-P99corresponds to Form I. The PXRD pattern of recrystallized Formula (1)from ethanol is depicted in FIG. 49. DSC of sample PP502-P99 revealed amelting onset at about 211° C. and an endothermic peak near 213° C.(with an enthalpy of fusion of 61.57 J/g) followed immediately by anexothermic degradation event.

TG-FTIR analysis of sample PP502-P99 revealed a mass loss of about 0.2%due to water loss and an additional mass loss of about 0.6% at 220° C.attributable to ethanol. A small amount of water and possibly a veryminor part of thermal degradation products is observed to begin uponmelting as heating continues thereafter to approximately 300° C.

The chemical identity and integrity of the recrystallized Formula (1)was confirmed by ¹H NMR spectroscopy. The triplet signal around 1.06 ppmis due to ethanol, and based on the integration of the split singleproton at 5.50/5.71 ppm, the ethanol content is calculated to be about0.054 equivalent which corresponds to 0.54%. No other residual solventappears to be present. The purity of the recrystallized Form I ofFormula (1) was confirmed by HPLC.

Particle size distribution testing was conducted using Fraunhofer laserdiffraction technique in n-heptane using a Malvern Mastersizer 2000.Values for the maximum particle size for a given percentage volume ofthe sample shown in Table 19 below. Furthermore, the particle sizedistribution function for the recrystallized base form indicates a peakat approximately 20 μm.

TABLE 19 Particle size distribution for recrystallized Form I free base.Sample x10 x50 (median) x90 P502-P99 3 μm 15 μm 37 μm

Hygroscopic behavior of the recrystallized Formula (1) (sample P502-P99)was measured using dynamic vapor sorption. The DVS results indicate thatthe recrystallized Formula (1) starts with an initial water content ofapproximately 0.25% at 50% RH, decreasing to about 0% at 0% RH, andreaching a maximum saturation of approximately 0.5% at a RH of about95%. DVS confirms that Formula (1) is not hygroscopic and the waterabsorption within the timeframe of the test is less than about 0.3% at95% RH. Moisture adsorption-desorption isotherms were prepared in asimilar manner as described above.

The recrystallized Formula (1) from ethanol was obtained as acrystalline material; examination by polarized optical microscopyrevealed that the material consists of particles ranging in sizes from afew microns, for the smaller particles, to about 100 μm for the largestparticles.

Example 9 Crystallization Optimization for Formula (1) Free Base Form I

Crystallization experiments were performed towards optimized productionof Formula (1) free base Form I. The starting material for thecrystallization experiments was recrystallized Formula (1) Form I. Thestudy was supplemented with additional crystallizations of the free basefrom a sample of crude oil.

The obtained products were characterized using powder X-ray diffraction(PXRD) or Raman spectroscopy to investigate crystalline form and byTG-FTIR or ¹H NMR or both to investigate residual solvent contents.Polarized microscopy images were recorded to determine particle size.

Acetone, ethanol, and 1-propanol are the most promising solvents for therecrystallization of Form I. Since crystalline Form I has a lowsolubility in many ICH class 3 solvents, the addition of potentiallyuseful co-solvents was explored. For example, ethanol, water, and aceticacid are solvents that may be used to increase the solubility of Form I,which is important in the design of a crystallization process thatmaximizes volume efficiency and yield.

Solubility data was collected for several solvent systems. Temperaturedependence of the Form I solubility was estimated for acetone, ethanol,ethanol-water 94:4 (v/v) and 1-propanol. Linear and non-linear coolingprofiles and various temperature cycling strategies were applied inorder to improve the quality of crystalline nature of Form I.

One method is based on crystallization of the maleate salt from a crudeoil wherein the crystalline maleate salt is neutralized with base andthe free base is extracted (presumably in amorphous form). Thereafter,the free base is crystallized from acetone and crystalline Form I(anhydrous form) is obtained. The resultant crystalline Form Iconsistently contain substantial amounts of residual solvent even whenPXRD patterns of all produced samples are identical.

For example, a sample of recrystallized Form I (sample PP502-P1)contains about 0.9% of acetone as determined by TG-FTIR. No mass loss isobservable below about 200° C.; however, heating thereafter results in arelease of acetone solvent along with melting of the solid form (meltingpoint of the solid form is approximately 215° C.). Prolonged drying atconventional drying temperatures does not necessarily efficiently reducethe residual solvents. However, recrystallization from other solvents(e.g., ethanol) have been shown to remove residual solvent from Form I.

Crystallization of an amorphous material after conversion of a salt tothe free base is fundamentally different from the process ofrecrystallization of a stable polymorphic form such as Form I. Form I ismuch less soluble than the amorphous form since it is typicallyrecovered after the extraction and evaporation of the solvent; however,solubility can change if the free base crystallizes spontaneously afterthe extraction step. Thought the specific difference in solubilitybetween the amorphous form and the stable crystalline form is not known,it ranges from a factor of 10 to 100.

In the current methodology, 100 mg/mL of stable Form I is preferablymixed into an ICH class 3 solvent or solvent mixture for the purposes ofrecrystallization. Solvent possibilities were narrowed by collectingdetailed solubility data for the most common solvents. Formula (1) isnot known to crystallize in different polymorphs, i.e., no othernon-solvated form was obtained from a crystallization experiment from asaturated solution.

The polymorphism study showed that Form I is stable and that this formis obtained consistently when the water activity was below the criticallimit for hydrate formation. A seeded process is recommended becauseseeding allows a better control of the crystallization process to obtaina more reproducible form, particle size and shape distribution. Thesamples in Table 20 were used in this study.

TABLE 20 Samples used in the optimization of the crystallization processfor Formula (1) free base Form I study. Sample name Batch No. Samplecode Form Formula (1), CS13-083 HB873-98 PP502-P1 Form I recrystallizedFormula (1), crude oil CS13-083 HB933-54-4 PP502-P61 Oil Formula (1),CS13-083 HB933-54-5 PP502-P67 Crystalline (CML 1476) maleate saltFormula (1), CS13-083, Am-1406 PP502-P62 Form I recrystallized

Example 9.1 Solubility by HPLC

The solubility of recrystallized Formula (1) Form I was tested invarious water-solvent mixtures and non-aqueous solvents. Completesolubility data generated for these and other solvent systems ispresented in Table 21 below.

TABLE 21 Solubility data for Form I. MEK refers to methyl ethyl ketoneand THF refers to tetrahydrofuran. Solubility Solubility Pure solvent(S, mg/mL) Solvent mixture (S, mg/mL) acetone, 25° C. 4.1 ethanol-water6.7 96:4, 0° C. acetone at RFT 10.0 ethanol-water 10.0 96:4, 25° C.ethyl acetate, 25° C. 1.5 ethanol-water 24.7 96:4, 60° C. ethanol, 5° C.3.6 ethanol-water 23.7 9:1, 25° C. ethanol, 25° C. 4.4 ethanol, 50° C.10.2 MEK, 25° C. 3.7 methanol, 25° C. 19.9 1-propanol, 5° C. 3.41-propanol, r.t. 4.7 1-propanol, 25° C. 4.0 1-propanol, 60° C. 14.42-propanol, 25° C. 1.3 THF, 25° C. 20.4

Acetone, ethanol, 96%-ethanol and 1-propanol were considered as the mostpromising solvent systems. Solubility in 96% ethanol is rather high atroom temperature (approximately 22° C.) and cooling to low temperaturewould be necessary to obtain good yields. Cooling below 0° C. was notexplored during the polymorphism study as crystallization at sub-zerotemperatures leads to hydrate formation. Though the presence of water inhigh-temperature co-solvent mixtures might lead to deteriorating Formula(1) stability (indicated by a red discoloration), water still serves asa useful co-solvent at low concentration levels from about 0.5 to 4%.

Example 9.2 Multimax Solubility Tests

Metastable zone width experiments were conducted in a Mettler-ToledoMultimax crystallization process optimization system equipped withturbidity probes, in order to demonstrate control of crystallization forForm I.

Acetone, ethanol, and ethanol-water (96:4) were selected as the solventsystems. Three different concentrations were selected for acetone andethanol; two different concentrations were selected for theethanol-water (96:4) solvent system. Solubility data obtained from theMultimax experiments were in agreement with previously obtained data;however, values from the Multimax experiment are slightly lower than theactual value due to the kinetic nature of the Multimax experiment. Thetemperature dependence of the solubility of the Formula (1) free baseForm I in ethanol and acetone is depicted in FIG. 48.

Ethanol and 1-propanol display similar solubility characteristics giventhat data points for the 1-propanol solvent appear to align well withthe curve fit for the ethanol data points. Moreover, because the boilingpoint of 1-propanol is 97° C. as compared to a boiling point of 78° C.for ethanol, 1-propanol is considered a viable alternative to ethanolresulting in a substantial increase of the volume efficiency and yield.

Without seeding, the crystallization experiments from cooling ofsupersaturated solutions did not lead to crystallization in any of theexamined solvents. As a consequence, the metastable zones in all testedsolvents are very wide. Therefore, seeding is mandatory to control thecrystallization process and is applied soon after oversaturation hasbeen achieved.

Example 9.3 Experimental Approach to Reduction of Residual SolventsExample 9.3.1 Temperature Cycling Experiments—Part One

Temperature cycling experiments were carried out to investigate theorigin of high residual solvent contents, which are hypothesized to bedue to solvate formation, or due to an inherent property of the freebase Form I to form “solvent inclusion” complexes, or due to crystaldefects that allow solvent inclusion. Polymorphism studies did not offerany positive proof for solvate formation for acetone, ethanol, ethylacetate and 1-propanol, though solvated forms were found with methanoland acetic acid. However, substantial amounts of residual solvent werefound for all of the mentioned solvents. Specifically, temperaturecycling experiments were conducted to test whether high residual solventcontents are due to crystal defects. The experiments were carried out infour different solvent systems: acetone, ethanol, 1-propanol and a 1:1mixture of methanol and TBME. Experiments with a T-cycling profile asshown in FIG. 50 were run between 25° C. and 50° C. for two days.

After 48 hours the suspensions were filtered, the obtained solids weredried under vacuum at 60° C. and tested by TG-FTIR and opticalmicroscopy. The TG-FTIR results are summarized in Table 22 and theoptical microscopy results are presented in FIG. 51. In all experiments,the solvent content was slightly reduced. In experiments with ethanolsolvent and methanol-TBME mixture, no acetone could be found by TG-FTIR.Optical microscopy shows that in the ethanol, acetone, and methanol-TBMEmixture experiments, (the exception is 1-propanol) very small particleswere obtained. Compared to the starting material of sample PP502-P1,particle size distribution shifts to smaller particles. The particles ofsamples PP502-P55 (in ethanol), PP502-P56 (in acetone) and PP502-P58 (inthe methanol-TBME mixture) have sizes of a few μm which border the limitof microscopic resolution. Stirring with a magnet bar might possiblyhave led to a milling effect over the two day cycling period. Theexception of sample PP502-P57 in 1-propanol might be due to irregularrotation of the magnet bars in the temperature controlled apparatus.

TABLE 22 TG-FTIR results from the temperature cycling experiments withFormula (1) free base Form I (containing approximately 0.8% of acetoneat the beginning of the experiment). Summarized Sample ConditionsTG-FTIR result PP502-P55 Form I in ethanol Mass loss 0.69%; likelywater, but not clearly attributable PP502-P56 Form I in acetone Massloss 0.58%; attributable to acetone PP502-P57 Form I in 1-propanol Massloss 0.55%; attributable to acetone PP502-P58 Form I in Mass loss 0.43%;likely water, methanol-TBME 1:1 but not clearly attributable

The results shown in Table 22 suggest a residual solvent reduction ofabout 30%; though this may be attributed to a milling effect rather thanto elimination of defect sites in crystalline particles. Therefore,wet-milling might be one way to reduce residual solvent content andethanol might be a good solvent in the wet-milling process.

Example 9.3.2 Temperature Cycling Experiments—Part Two

Additional temperature cycling experiments were carried out in acetoneand ethanol using two different temperature cycling programs. Differentfrom the previous experiments, these experiments were carried out in theMultimax crystallization system. The first program consisted of fourcycles with a duration of 12 hours each (cooling rate of 4 K/h) and thesecond program consisted of eight cycles with a duration of 6 hours each(cooling rate of 7.5 K/h). Both temperature cycling programs covered atemperature range of from about 23° C. to about 53° C. and ran for 48hours and subsequently a cooling to 3° C. at the end of the experiment.Turbidity in the system was recorded with a turbidity probe (not shown).In all experiments, concentration of solid was about 50 mg/mL ofsolvent, i.e., based on the solubility results for ethanol, about 10% ofthe solid is dissolved at the beginning of the cycle and about 20% ofthe solid is dissolved at the high temperature end of the cycle.

Aliquots were taken and analyzed by PXRD and TG-FTIR after 24 hours andafter 48 hours. After the samples were cooled to 3° C., the samples wereagain isolated and analyzed by PXRD, optical microscopy, and TG-FTIR. Asummary of the results from the TG-FTIR is given in Table 23. PXRDshowed that all samples were obtained as Form I. Although a slightreduction in the acetone content is observable, TG-FTIR mass lossesstill suggest an acetone content of about 0.7%.

TABLE 23 Results of equilibration experiments with temperature cycling.Sample Solvent Temperature Program TG-FTIR result PP502-P71 ethanol 4cycles, 12 hours each PP502-P71A ~0.78% acetone. PP502-P71B ~0.66%acetone. PP502-P71 (end) ~0.68% acetone. PP502-P72 acetone 4 cycles, 12hours each PP502-P72A ~0.68% acetone. PP502-P72B ~0.67% acetone.PP502-P72 ~0.71% acetone. PP502-P73 ethanol 8 cycles, 6 hours eachPP502-P73A ~0.65% acetone. PP502-P73B ~0.77% acetone. PP502-P73 ~0.75%acetone. PP502-P74 acetone 8 cycles, 6 hours each PP502-P74A ~0.67%acetone. PP502-P74B ~0.70% acetone. PP502-P72 ~0.72% acetone.

Example 9.3.3 Crystallization by Slow Cooling of Saturated Solutions

Two preliminary experiments were carried out in ethanol-water 96:4 (v/v)and in 1-propanol. Optical microscopy of samples PP502-P59 and PP502-P60(not shown) revealed that particle sizes were small and the residualsolvent content was slightly reduced with respect to the startingmaterial for sample PP502-P60 in 96% ethanol, but not for samplePP502-P59 in 1-propanol.

Additional slow cooling crystallization experiments starting from clearsolutions were carried out in a Multimax apparatus with a 1-propanol/H₂O98:2 mixture and an ethanol/H₂O ˜97:3 mixture using two differentcooling profiles. For both profiles, Form I was dissolved at 85° C.; thesolution was cooled to 75° C., seeded with Form I, and then cooled downto 6° C. for the 1-propanol/H₂O mixture or 4° C. for the ethanol/H₂Omixture. The first program (for samples PP502-P75 and PP502-P76)consisted of a stepwise cooling with integrated temperature cycling(cooling rate of 5 K/h) while the second program (for samples PP502-P77and PP502-P78) consisted of a continuous, non-linear cooling profile(starting cooling rate of 0.5 K/h and stepwise increase at lowertemperatures to a final cooling rate of 16 K/h). The cooling phase ofboth crystallization experiments was about 22 hours.

The turbidity of the samples was measured in-situ in order to monitorthe crystallization process. With the exception of PP502-P78, a slow andgradual increase of turbidity was observed during the cooling process,which is an indication of a controlled crystallization process.Turbidity analysis for sample PP502-P78 reflected that seed crystalswere dissolved because the solution was not saturated when seed crystalswere added. The samples were worked up and examined by PXRD, opticalmicroscopy and TG-FTIR. Powder X-ray diffraction of sample PP502-P78resulted in a pattern similar to the pattern for the amorphous form andshowed that the PP502-P78 sample did not crystallize. The PXRD patternsof all other samples (PP502-P75 to PP502-P77) correspond to Form I.

The optical microscopy images are presented in FIG. 52 and a summary ofthe TG-FTIR results are shown in Table 24. Whereas optical microscopyshowed that highly crystalline materials were obtained with particlesizes of up to about 100 μm, TG-FTIR results reflect that even a slow,gradual cooling crystallization does not result in solid materialwithout enclosed solvent residues, where solvent contents were typicallyabout 1% or slightly higher.

TABLE 24 Results of cooling crystallization experiments. Sample SolventCooling profile TG-FTIR result PP502-P75 2% H₂O in 1-propanol Stepwisecooling with 1.09% 1-propanol temperature cycling PP502-P76 3% H₂O inethanol Stepwise cooling with 0.94% ethanol temperature cyclingPP502-P77 2% H₂O in 1-propanol Slow continuous, non- 1.34% 1-propanollinear cooling PP502-P78 3% H₂O in ethanol Slow continuous, non- ~3%mass loss due to water linear cooling

Example 9.3.4 The Effect of Wet-Milling

Whereas the results from the first series of temperature cyclingexperiments (example 8.3.1 above) suggested that the solvent contentscan potentially be reduced by crystal ripening in suspensionexperiments, results from the second series of temperature cyclingexperiments (example 8.3.2 above) failed to confirm this result. Becauseoptical microscopy showed that particles in experiments PP502-P55through PP502-P58 were generally small and particles from experimentsPP502-P71 through PP502-P74 were large crystals, it was suspected thatthe use of different magnetic stir bars had caused a milling effect inthe former case while more gentle stirring (and slower cooling rates) inthe latter case allowed the formation of larger crystals.

Here, a suspension of sample PP502-P77 containing about 1.3% of1-propanol (see Table 24 above) was stirred with a fairly large magneticstirrer at about 1000 rpm for four days at room temperature. Opticalmicroscopy images showing the comparison between stirring processesrevealed that particle size distribution changes with stirringconditions. Whereas sample PP502-P77 contained particles having sizes upto about 100 μm, the slurried sample PP502-P88 does not appear tocontain any particles greater than 10 μm.

Due to small particle sizes, filtration of sample PP502-P88 was slow.Investigation of PP502-P88 by ¹H NMR showed that the sample contains0.32% 1-propanol and 0.07% ethanol after drying at 60° C. under vacuumfor several hours. This experiment confirms that the residual solventsin samples with small particle sizes below about 10 μm may be reduced tobelow the ICH limits. The result also suggests that 1-propanol waspartially replaced by ethanol. As a consequence, if wet-milling isconsidered, a milling-solvent screen can be employed if ethanol is notsuitable. Where water leads to a partial formation of the hydrate, drymilling is not recommended because this leads to partial amorphizationof the compound.

Example 10 Crystal Structure of Form I of the Formula (1) Free Base

A study of the crystal structure of Formula (1) free base Form I wasperformed. Intensity data were collected at 173 K, using Cu radiation(λ=1.54184 Å), on an Oxford Diffraction Gemini-R Ultra diffractometeroperated by the CrysAlis software (Agilent Technologies, 2012, Yarnton,England; CrysAlis CCD and CrysAlis RED, 2003). The data were correctedfor absorption effects by means of comparison of equivalent reflections.The structures were solved with the direct methods procedure implementedin SHELXT and refined by full-matrix least squares on F² usingSHELXL-2014, as described in Sheldrick, Acta Cryst. 2008, A64, 112-122.

Non-hydrogen atoms were located in difference maps and refinedanisotropically.

Hydrogen atoms of the main component were located in difference maps andthose bonded to carbon atoms were fixed in idealized positions and theirthermal displacement parameters were set to 1.2 U_(eq) (CH and CH₂) or1.5 U_(eq) (CH₃ groups) of the parent C atom. The positions of NHhydrogen atoms were refined with N—H distances restrained to 0.86(2) Åand the U_(iso) parameters were refined freely.

The results of the crystallographic experiments are summarized in Table25.

TABLE 25 Crystal data and structure refinement for Form I.Identification code 14thg420_PP502-P703 Empirical formula C₂₆H₂₃N₇O₂Formula weight  465.51 Temperature 173(2) K Wavelength 1.54184 Å (Curadiation) Crystal system Triclinic Space group P1 Unit cell dimensionsa = 8.0630(6) Å α = 85.841(5)° b = 10.2949(6) Å β = 75.798(6)° c =14.2825(8) Å γ = 82.331(5)° Volume 1138.07(13) Å³ Z   2 Density(calculated) 1.358 Mg/m³ Absorption coefficient 0.733 mm⁻¹ F(000)  488Crystal block; colorless Crystal size 0.150 × 0.100 × 0.050 mm³ Thetarange for data collection 3.2-68.2° Index ranges −9 ≦ h ≦ 9, −12 ≦ k ≦12, −17 ≦ l ≦ 17 Reflections collected 20885 Independent reflections7763 [R_(int) = 0.0559] Completeness to theta = 67.7° 99.1% Absorptioncorrection Semi-empirical from equivalents Max. and min. transmission1.00000 and 0.80527 Refinement method Full-matrix least-squares on F²Data/restraints/parameters 7763/9/657 Goodness-of-fit on F²   1.024Final R indices [F² > 2σ(F²)] R1 = 0.0530, wR2 = 0.1351 R indices (alldata) R1 = 0.0645, wR2 = 0.1477 Absolute structure parameter −0.1(3)(Flack) Extinction coefficient none Largest diff. peak and hole 0.380and −0.227 e Å⁻³

The asymmetric unit of the Form I crystal structure contains twomolecules, denoted A and B (FIG. 53), which differ fundamentally intheir conformation. The absolute configuration at both C9 and C9′ wasestablished as the (S)-configuration by anomalous-dispersion effects.The Flack x parameters, −0.1(3), was determined using 2602 quotients[(I+)−(I−)]/[(I+)+(I−)], as described in Parsons, et al., Acta Cryst.2013, B69, 249-259). The hydrogen atoms of the NH₂ and NH groups werelocated in difference maps. They were refined with the N—H distancesbeing restrained to 0.86(2) Å (which results in nine distancerestraints), and their U_(iso) parameters were refined freely. Thecrystal structure contains classical N—H • • • O and N—H • • • N bonds(listed in Table 26) which involve the NH₂ groups of N19 and N19′ asH-bond donor sites. By contrast, neither of the NH groups (N25 and N25′)is engaged in a classical N—H • • • A interaction.

TABLE 26 Hydrogen bonds in the Form I crystal structure. D-H•••A d(D-H)(Å) d(H•••A) (Å) d(D•••A) (Å) DHA angle (Å) N(19)-H(19A)•••N(15′)#1 0.88(3) 2.23 (3) 3.082 (7) 163 (5) N(19)-H(19B)•••O(35′) 0.88 (3) 2.34 (5)2.997 (6) 132 (5) N(19′)-H(19C)•••N(15)#2 0.88 (3) 2.19 (3) 3.051 (7)168 (5) N(19′)-H(19D)•••O(35) 0.86 (3) 2.31 (4) 2.961 (6) 133 (4)Symmetry transformations used to generate equivalent atoms:#1 x−1,y+1,z #2 x+1,y−1,z

Additional results, including atomic coordinates for the Form I crystalstructure, are given in Table 27, Table 28, and Table 29.

TABLE 27 Atomic coordinates (×10⁴), equivalent isotropic displacementparameters (Å² × 10³) and site occupancy factors for heavy atoms in theForm I crystal structure. U_(eq) is defined as one third of the trace ofthe orthogonalized U_(ij) tensor. Atom x y z U_(eq) s.o.f. C(1)−5031(12) 4749(10) 11713(8) 102(4)  1 C(2) −4155(7) 5851(6) 11213(5)55(1) 1 C(3) −3457(6) 6761(5) 10816(4) 44(1) 1 C(4) −2490(6) 7774(5)10254(4) 38(1) 1 N(5) −815(5) 7611(4) 10249(3) 34(1) 1 C(6) 48(7)6576(5) 10786(5) 47(1) 1 C(7) 1914(7) 6819(6) 10537(5) 50(1) 1 C(8)1921(6) 8234(5) 10125(4) 42(1) 1 C(9) 384(6) 8486(5) 9660(4) 35(1) 1C(10) 869(6) 8113(4) 8620(4) 32(1) 1 N(11) 1834(5) 7013(4) 8313(3) 34(1)1 C(12) 1941(6) 6994(4) 7340(4) 32(1) 1 C(13) 1042(6) 8128(5) 7044(4)31(1) 1 C(14) 864(6) 8767(4) 6143(4) 32(1) 1 N(15) 42(5) 9955(4) 6099(3)40(1) 1 C(16) −699(7) 10570(5) 6961(4) 41(1) 1 C(17) −566(6) 10076(5)7829(4) 36(1) 1 N(18) 349(5) 8832(4) 7882(3) 31(1) 1 N(19) 1578(6)8135(4) 5299(3) 37(1) 1 C(20) 2983(6) 5898(4) 6779(4) 32(1) 1 C(21)2317(6) 5168(5) 6203(4) 39(1) 1 C(22) 3348(7) 4157(5) 5677(4) 42(1) 1C(23) 5067(6) 3866(4) 5712(4) 34(1) 1 C(24) 6328(7) 2859(5) 5117(4)37(1) 1 N(25) 5808(6) 2362(4) 4391(4) 42(1) 1 C(26) 6841(7) 1525(5)3677(4) 39(1) 1 N(27) 6489(6) 1771(5) 2817(4) 49(1) 1 C(28) 7406(8)1006(7) 2100(5) 62(2) 1 C(29) 8617(8) −12(8) 2228(6) 68(2) 1 C(30)8947(8) −248(6) 3131(6) 60(2) 1 C(31) 8054(6) 530(5) 3885(5) 45(1) 1C(32) 5713(6) 4562(5) 6308(4) 36(1) 1 C(33) 4676(6) 5560(5) 6840(4)35(1) 1 O(34) −3169(5) 8677(4) 9817(3) 50(1) 1 O(35) 7747(5) 2531(4)5270(3) 56(1) 1 C(1′) 8013(7) 5676(6) −1723(5) 54(2) 1 C(2′) 8922(7)4542(5) −1320(5) 48(1) 1 C(3′) 9675(6) 3596(5) −994(4) 44(1) 1 C(4′)10504(6) 2521(5) −495(4) 38(1) 1 N(5′) 12205(5) 2548(4) −578(3) 34(1) 1C(6′) 13250(7) 3509(5) −1178(5) 46(1) 1 C(7′) 14868(8) 3325(6) −838(6)63(2) 1 C(8′) 15030(7) 1917(5) −430(4) 46(1) 1 C(9′) 13158(6) 1680(5)19(4) 35(1) 1 C(10′) 12527(6) 2046(4) 1057(4) 30(1) 1 N(11′) 11591(5)3147(4) 1374(3) 34(1) 1 C(12′) 11338(6) 3088(5) 2362(4) 32(1) 1 C(13′)12134(5) 1901(4) 2658(4) 29(1) 1 C(14′) 12207(6) 1197(5) 3554(4) 33(1) 1N(15′) 12982(5) 0(4) 3586(3) 39(1) 1 C(16′) 13775(7) −571(5) 2716(4)40(1) 1 C(17′) 13765(6) −14(5) 1835(4) 34(1) 1 N(18′) 12895(5) 1245(4)1806(3) 31(1) 1 N(19′) 11457(6) 1795(4) 4404(3) 38(1) 1 C(20′) 10255(6)4156(5) 2948(4) 32(1) 1 C(21′) 10816(7) 4799(5) 3611(5) 43(1) 1 C(22′)9761(7) 5791(5) 4148(4) 42(1) 1 C(23′) 8077(6) 6141(5) 4049(4) 35(1) 1C(24′) 6789(6) 7134(5) 4630(4) 36(1) 1 N(25′) 7299(5) 7695(4) 5342(3)39(1) 1 C(26′) 6224(6) 8530(5) 6041(4) 36(1) 1 N(27′) 6563(6) 8331(4)6920(4) 43(1) 1 C(28′) 5595(7) 9096(6) 7612(5) 48(1) 1 C(29′) 4311(7)10047(6) 7481(5) 51(2) 1 C(30′) 3991(7) 10254(5) 6562(5) 47(1) 1 C(31′)4960(6) 9479(5) 5821(5) 41(1) 1 C(32′) 7535(7) 5502(5) 3372(4) 39(1) 1C(33′) 8588(6) 4532(5) 2832(4) 35(1) 1 O(34′) 9721(5) 1648(3) −30(3)49(1) 1 O(35′) 5357(5) 7389(4) 4493(3) 52(1) 1

TABLE 28 Bond lengths (Å) and angles (°) for the Form I crystalstructure. C(1)-C(2)  1.465(9) C(1)-H(1A)  0.98 C(1)-H(1B)  0.98C(1)-H(1C)  0.98 C(2)-C(3)  1.193(8) C(3)-C(4)  1.459(8) C(4)-O(34) 1.225(6) C(4)-N(5)  1.337(6) N(5)-C(6)  1.468(7) N(5)-C(9)  1.475(6)C(6)-C(7)  1.509(8) C(6)-H(6A)  0.99 C(6)-H(6B)  0.99 C(7)-C(8) 1.532(8) C(7)-H(7A)  0.99 C(7)-H(7B)  0.99 C(8)-C(9)  1.528(7)C(8)-H(8A)  0.99 C(8)-H(8B)  0.99 C(9)-C(10)  1.505(7) C(9)-H(9)  1C(10)-N(11)  1.323(6) C(10)-N(18)  1.363(7) N(11)-C(12)  1.372(7)C(12)-C(13)  1.386(7) C(12)-C(20)  1.472(7) C(13)-N(18)  1.399(6)C(13)-C(14)  1.432(7) C(14)-N(15)  1.316(6) C(14)-N(19)  1.372(7)N(15)-C(16)  1.386(7) C(16)-C(17)  1.330(8) C(16)-H(16)  0.95C(17)-N(18)  1.397(6) C(17)-H(17)  0.95 N(19)-H(19A)  0.88(3)N(19)-H(19B)  0.88(3) C(20)-C(33)  1.385(7) C(20)-C(21)  1.395(7)C(21)-C(22)  1.384(7) C(21)-H(21)  0.95 C(22)-C(23)  1.390(7)C(22)-H(22)  0.95 C(23)-C(32)  1.385(7) C(23)-C(24)  1.501(7)C(24)-O(35)  1.217(6) C(24)-N(25)  1.363(7) N(25)-C(26)  1.413(7)N(25)-H(25)  0.86(3) C(26)-N(27)  1.327(8) C(26)-C(31)  1.387(8)N(27)-C(28)  1.343(8) C(28)-C(29)  1.368(11) C(28)-H(28)  0.95C(29)-C(30)  1.377(11) C(29)-H(29)  0.95 C(30)-C(31)  1.383(8)C(30)-H(30)  0.95 C(31)-H(31)  0.95 C(32)-C(33)  1.380(7) C(32)-H(32) 0.95 C(33)-H(33)  0.95 C(1′)-C(2′)  1.453(8) C(1′)-H(1′1)  0.98C(1′)-H(1′2)  0.98 C(1′)-H(1′3)  0.98 C(2′)-C(3′)  1.204(8) C(3′)-C(4′) 1.449(8) C(4′)-O(34′)  1.228(6) C(4′)-N(5′)  1.351(6) N(5′)-C(9′) 1.465(6) N(5′)-C(6′)  1.472(7) C(6′)-C(7′)  1.485(9) C(6′)-H(6′1)  0.99C(6′)-H(6′2)  0.99 C(7′)-C(8′)  1.523(9) C(7′)-H(7′1)  0.99 C(7′)-H(7′2) 0.99 C(8′)-C(9′)  1.532(7) C(8′)-H(8′1)  0.99 C(8′)-H(8′2)  0.99C(9′)-C(10′)  1.502(7) C(9′)-H(9′)  1 C(10′)-N(11′)  1.318(6)C(10′)-N(18′)  1.370(6) N(11′)-C(12′)  1.373(7) C(12′)-C(13′)  1.390(7)C(12′)-C(20′)  1.480(7) C(13′)-N(18′)  1.398(6) C(13′)-C(14′)  1.434(7)C(14′)-N(15′)  1.308(6) C(14′)-N(19′)  1.366(7) N(15′)-C(16′)  1.383(7)C(16′)-C(17′)  1.346(8) C(16′)-H(16′)  0.95 C(17′)-N(18′)  1.392(6)C(17′)-H(17′)  0.95 N(19′)-H(19C)  0.88(3) N(19′)-H(19D)  0.86(3)C(20′)-C(21′)  1.386(7) C(20′)-C(33′)  1.394(7) C(21′)-C(22′)  1.385(8)C(21′)-H(21′)  0.95 C(22′)-C(23′)  1.396(7) C(22′)-H(22′)  0.95C(23′)-C(32′)  1.391(7) C(23′)-C(24′)  1.493(7) C(24′)-O(35′)  1.209(6)C(24′)-N(25′)  1.376(7) N(25′)-C(26′)  1.409(7) N(25′)-H(25′)  0.85(3)C(26′)-N(27′)  1.345(7) C(26′)-C(31′)  1.392(7) N(27′)-C(28′)  1.334(7)C(28′)-C(29′)  1.366(9) C(28′)-H(28′)  0.95 C(29′)-C(30′)  1.395(10)C(29′)-H(29′)  0.95 C(30′)-C(31′)  1.385(8) C(30′)-H(30′)  0.95C(31′)-H(31′)  0.95 C(32′)-C(33′)  1.370(7) C(32′)-H(32′)  0.95C(33′)-H(33′)  0.95 C(2)-C(1)-H(1A) 109.5 C(2)-C(1)-H(1B) 109.5H(1A)-C(1)-H(1B) 109.5 C(2)-C(1)-H(1C) 109.5 H(1A)-C(1)-H(1C) 109.5H(1B)-C(1)-H(1C) 109.5 C(3)-C(2)-C(1) 179.0(9) C(2)-C(3)-C(4) 173.8(6)O(34)-C(4)-N(5) 123.7(5) O(34)-C(4)-C(3) 122.1(4) N(5)-C(4)-C(3)114.1(4) C(4)-N(5)-C(6) 125.8(4) C(4)-N(5)-C(9) 121.5(4) C(6)-N(5)-C(9)112.6(4) N(5)-C(6)-C(7) 105.2(5) N(5)-C(6)-H(6A) 110.7 C(7)-C(6)-H(6A)110.7 N(5)-C(6)-H(6B) 110.7 C(7)-C(6)-H(6B) 110.7 H(6A)-C(6)-H(6B) 108.8C(6)-C(7)-C(8) 105.3(4) C(6)-C(7)-H(7A) 110.7 C(8)-C(7)-H(7A) 110.7C(6)-C(7)-H(7B) 110.7 C(8)-C(7)-H(7B) 110.7 H(7A)-C(7)-H(7B) 108.8C(9)-C(8)-C(7) 105.3(4) C(9)-C(8)-H(8A) 110.7 C(7)-C(8)-H(8A) 110.7C(9)-C(8)-H(8B) 110.7 C(7)-C(8)-H(8B) 110.7 H(8A)-C(8)-H(8B) 108.8N(5)-C(9)-C(10) 110.3(4) N(5)-C(9)-C(8) 102.4(4) C(10)-C(9)-C(8)111.9(4) N(5)-C(9)-H(9) 110.7 C(10)-C(9)-H(9) 110.7 C(8)-C(9)-H(9) 110.7N(11)-C(10)-N(18) 111.2(4) N(11)-C(10)-C(9) 123.6(5) N(18)-C(10)-C(9)125.1(4) C(10)-N(11)-C(12) 106.8(4) N(11)-C(12)-C(13) 109.6(4)N(11)-C(12)-C(20) 119.8(4) C(13)-C(12)-C(20) 130.5(5) C(12)-C(13)-N(18)105.4(4) C(12)-C(13)-C(14) 136.7(5) N(18)-C(13)-C(14) 117.4(4)N(15)-C(14)-N(19) 118.7(5) N(15)-C(14)-C(13) 121.8(5) N(19)-C(14)-C(13)119.5(4) C(14)-N(15)-C(16) 117.9(5) C(17)-C(16)-N(15) 124.8(5)C(17)-C(16)-H(16) 117.6 N(15)-C(16)-H(16) 117.6 C(16)-C(17)-N(18)117.7(5) C(16)-C(17)-H(17) 121.1 N(18)-C(17)-H(17) 121.1C(10)-N(18)-C(17) 132.6(4) C(10)-N(18)-C(13) 107.0(4) C(17)-N(18)-C(13)120.2(4) C(14)-N(19)-H(19A) 109(4) C(14)-N(19)-H(19B) 115(4)H(19A)-N(19)-H(19B) 116(6) C(33)-C(20)-C(21) 118.6(4) C(33)-C(20)-C(12)119.0(4) C(21)-C(20)-C(12) 122.3(4) C(22)-C(21)-C(20) 120.5(4)C(22)-C(21)-H(21) 119.8 C(20)-C(21)-H(21) 119.8 C(21)-C(22)-C(23)120.2(5) C(21)-C(22)-H(22) 119.9 C(23)-C(22)-H(22) 119.9C(32)-C(23)-C(22) 119.3(4) C(32)-C(23)-C(24) 116.1(4) C(22)-C(23)-C(24)124.5(4) O(35)-C(24)-N(25) 122.5(5) O(35)-C(24)-C(23) 121.1(4)N(25)-C(24)-C(23) 116.4(4) C(24)-N(25)-C(26) 126.1(4) C(24)-N(25)-H(25)114(4) C(26)-N(25)-H(25) 117(4) N(27)-C(26)-C(31) 124.8(5)N(27)-C(26)-N(25) 113.2(5) C(31)-C(26)-N(25) 122.1(5) C(26)-N(27)-C(28)116.7(6) N(27)-C(28)-C(29) 123.4(7) N(27)-C(28)-H(28) 118.3C(29)-C(28)-H(28) 118.3 C(28)-C(29)-C(30) 118.5(6) C(28)-C(29)-H(29)120.7 C(30)-C(29)-H(29) 120.7 C(29)-C(30)-C(31) 120.0(6)C(29)-C(30)-H(30) 120 C(31)-C(30)-H(30) 120 C(30)-C(31)-C(26) 116.6(6)C(30)-C(31)-H(31) 121.7 C(26)-C(31)-H(31) 121.7 C(33)-C(32)-C(23)120.3(4) C(33)-C(32)-H(32) 119.9 C(23)-C(32)-H(32) 119.9C(32)-C(33)-C(20) 121.0(4) C(32)-C(33)-H(33) 119.5 C(20)-C(33)-H(33)119.5 C(2′)-C(1′)-H(1′1) 109.5 C(2′)-C(1′)-H(1′2) 109.5H(1′1)-C(1′)-H(1′2) 109.5 C(2′)-C(1′)-H(1′3) 109.5 H(1′1)-C(1′)-H(1′3)109.5 H(1′2)-C(1′)-H(1′3) 109.5 C(3′)-C(2′)-C(1′) 179.4(6)C(2′)-C(3′)-C(4′) 173.5(6) O(34′)-C(4′)-N(5′) 122.3(5)O(34′)-C(4′)-C(3′) 122.6(4) N(5′)-C(4′)-C(3′) 115.1(5) C(4′)-N(5′)-C(9′)121.7(4) C(4′)-N(5′)-C(6′) 125.2(4) C(9′)-N(5′)-C(6′) 112.8(4)N(5′)-C(6′)-C(7′) 103.7(5) N(5′)-C(6′)-H(6′1) 111 C(7′)-C(6′)-H(6′1) 111N(5′)-C(6′)-H(6′2) 111 C(7′)-C(6′)-H(6′2) 111 H(6′1)-C(6′)-H(6′2) 109C(6′)-C(7′)-C(8′) 105.8(5) C(6′)-C(7′)-H(7′1) 110.6 C(8′)-C(7′)-H(7′1)110.6 C(6′)-C(7′)-H(7′2) 110.6 C(8′)-C(7′)-H(7′2) 110.6H(7′1)-C(7′)-H(7′2) 108.7 C(7′)-C(8′)-C(9′) 103.5(4) C(7′)-C(8′)-H(8′1)111.1 C(9′)-C(8′)-H(8′1) 111.1 C(7′)-C(8′)-H(8′2) 111.1C(9′)-C(8′)-H(8′2) 111.1 H(8′1)-C(8′)-H(8′2) 109 N(5′)-C(9′)-C(10′)109.5(4) N(5′)-C(9′)-C(8′) 102.6(4) C(10′)-C(9′)-C(8′) 113.8(4)N(5′)-C(9′)-H(9′) 110.2 C(10′)-C(9′)-H(9′) 110.2 C(8′)-C(9′)-H(9′) 110.2N(11′)-C(10′)-N(18′) 111.1(4) N(11′)-C(10′)-C(9′) 126.2(4)N(18′)-C(10′)-C(9′) 122.6(4) C(10′)-N(11′)-C(12′) 107.0(4)N(11′)-C(12′)-C(13′) 109.7(4) N(11′)-C(12′)-C(20′) 120.5(4)C(13′)-C(12′)-C(20′) 129.6(5) C(12′)-C(13′)-N(18′) 105.1(4)C(12′)-C(13′)-C(14′) 137.3(4) N(18′)-C(13′)-C(14′) 117.3(4)N(15′)-C(14′)-N(19′) 118.7(5) N(15′)-C(14′)-C(13′) 122.1(4)N(19′)-C(14′)-C(13′) 119.2(4) C(14′)-N(15′)-C(16′) 117.7(5)C(17′)-C(16′)-N(15′) 125.3(4) C(17′)-C(16′)-H(16′) 117.3N(15′)-C(16′)-H(16′) 117.3 C(16′)-C(17′)-N(18′) 116.7(5)C(16′)-C(17′)-H(17′) 121.6 N(18′)-C(17′)-H(17′) 121.6C(10′)-N(18′)-C(17′) 132.0(4) C(10′)-N(18′)-C(13′) 107.1(3)C(17′)-N(18′)-C(13′) 120.8(4) C(14′)-N(19′)-H(19C) 112(4)C(14′)-N(19′)-H(19D) 121(3) H(19C)-N(19′)-H(19D) 110(5)C(21′)-C(20′)-C(33′) 118.3(4) C(21′)-C(20′)-C(12′) 123.1(4)C(33′)-C(20′)-C(12′) 118.6(4) C(22′)-C(21′)-C(20′) 121.3(5)C(22′)-C(21′)-H(21′) 119.3 C(20′)-C(21′)-H(21′) 119.3C(21′)-C(22′)-C(23′) 120.2(5) C(21′)-C(22′)-H(22′) 119.9C(23′)-C(22′)-H(22′) 119.9 C(32′)-C(23′)-C(22′) 117.9(5)C(32′)-C(23′)-C(24′) 117.0(4) C(22′)-C(23′)-C(24′) 125.1(4)O(35′)-C(24′)-N(25′) 122.5(4) O(35′)-C(24′)-C(23′) 121.0(4)N(25′)-C(24′)-C(23′) 116.5(4) C(24′)-N(25′)-C(26′) 125.5(4)C(24′)-N(25′)-H(25′) 123(4) C(26′)-N(25′)-H(25′) 111(4)N(27′)-C(26′)-C(31′) 124.1(5) N(27′)-C(26′)-N(25′) 113.5(4)C(31′)-C(26′)-N(25′) 122.3(5) C(28′)-N(27′)-C(26′) 116.5(5)N(27′)-C(28′)-C(29′) 124.4(6) N(27′)-C(28′)-H(28′) 117.8C(29′)-C(28′)-H(28′) 117.8 C(28′)-C(29′)-C(30′) 118.4(5)C(28′)-C(29′)-H(29′) 120.8 C(30′)-C(29′)-H(29′) 120.8C(31′)-C(30′)-C(29′) 119.2(5) C(31′)-C(30′)-H(30′) 120.4C(29′)-C(30′)-H(30′) 120.4 C(30′)-C(31′)-C(26′) 117.4(6)C(30′)-C(31′)-H(31′) 121.3 C(26′)-C(31′)-H(31′) 121.3C(33′)-C(32′)-C(23′) 121.8(5) C(33′)-C(32′)-H(32′) 119.1C(23′)-C(32′)-H(32′) 119.1 C(32′)-C(33′)-C(20′) 120.4(5)C(32′)-C(33′)-H(33′) 119.8 C(20′)-C(33′)-H(33′) 119.8

TABLE 29 Hydrogen coordinates [×10⁴] and isotropic displacementparameters [Å² × 10³] for the Form I crystal structure. Atom x y zU_(eq) s.o.f. H(1A) −6178 5079 12092 153 1 H(1B) −4356 4281 12145 153 1H(1C) −5145 4148 11237 153 1 H(6A) −68 5698 10586 57 1 H(6B) −452 663211490 57 1 H(7A) 2615 6200 10053 60 1 H(7B) 2383 6712 11121 60 1 H(8A)1793 8851 10645 51 1 H(8B) 3009 8338 9638 51 1 H(9) −136 9423 9718 42 1H(16) −1351 11404 6928 49 1 H(17) −1075 10553 8395 43 1 H(19A) 1750 (70)8720 (40) 4820 (30) 38 (15) 1 H(19B) 2460 (60) 7530 (50) 5320 (40) 41(15) 1 H(21) 1146 5365 6172 47 1 H(22) 2881 3660 5290 50 1 H(25) 4900(50) 2800 (50) 4260 (40) 42 (15) 1 H(28) 7204 1182 1473 74 1 H(29) 9216−545 1706 81 1 H(30) 9788 −946 3235 72 1 H(31) 8260 391 4513 54 1 H(32)6876 4351 6352 44 1 H(33) 5129 6022 7253 42 1 H(1′1) 8252 5619 −2426 811 H(1′2) 8403 6479 −1568 81 1 H(1′3) 6772 5696 −1446 81 1 H(6′1) 126644414 −1075 55 1 H(6′2) 13481 3328 −1872 55 1 H(7′1) 14809 3950 −333 75 1H(7′2) 15868 3464 −1381 75 1 H(8′1) 15714 1821 65 55 1 H(8′2) 15577 1299−948 55 1 H(9′) 13007 745 −46 42 1 H(16′) 14375 −1427 2743 48 1 H(17′)14324 −458 1261 41 1 H(19C) 11220 (70)  1220 (50) 4890 (30) 41 (16) 1H(19D) 10600 (50)  2400 (40) 4440 (40) 22 (12) 1 H(21′) 11948 4553 369952 1 H(22′) 10185 6236 4584 50 1 H(25′) 8300 (50) 7510 (50) 5450 (50) 41(15) 1 H(28′) 5814 8970 8239 57 1 H(29′) 3650 10556 8004 62 1 H(30′)3120 10920 6446 56 1 H(31′) 4770 9590 5188 49 1 H(32′) 6404 5746 3281 471 H(33′) 8180 4113 2376 42 1

The PXRD pattern of Form I is compared to the pattern simulated usingthe crystal structure of Form I in FIG. 54. The patterns show agreementthat indicates that the single crystal is representative of the samecrystalline phase as Form I.

Example 11 Crystal Structure of Form III of the Formula (1) Free Base

A study of the crystal structure of Formula (1) free base Form III wasalso performed. The experimental methods are as described in Example 9,with the exception of the following. Water hydrogen atoms werepositioned in assumed ideal positions and refined with O—H and H • • • Hdistances restrained to 0.86(1) Å and 1.51(2) Å, respectively, and theirthermal displacement parameters were set to 1.2 U_(eq) of the parent Oatom. The O4W/O4W′ water molecule was found to be disordered over twopositions with relative occupancies of 0.85 and 0.15. The hydrogen atomsof the O4W/O4W′ molecule could not be determined, but they are includedin the molecular formula. The rings N5-C6-C7-C8-C9 of molecule A andN5′-C6′-C7′-C8′-C9′ of molecule B are each disordered over twopositions, with the relative occupancies for their major components of0.70 and 0.61, respectively. Distance restraints were applied onequivalent bond and non-bond distances of the disorder component. Theopposite atoms of disorder fragments were refined with equal anisotropicdisplacement parameters.

The results of the crystallographic experiments are summarized in Table30.

TABLE 30 Crystal data and structure refinement for Form III.Identification code 14thg422_PP502-Sol-DMF-H2O Empirical formulaC₂₆H₂₇N₇O₄ Moiety formula C₂₆H₂₃N₇O₂•2H₂O Formula weight 501.54Temperature 293 (2) K Wavelength 1.54184 Å (Cu radiation) Crystal systemMonoclinic Space group P2₁ Unit cell dimensions a = 8.3932 (6) Å α = 90°b = 21.1721 (11) Å β = 94.586 (5)° c = 14.1298 (7) Å γ = 90° Volume2502.9 (3) Å³ Z 4 Density (calculated) 1.331 Mg/m³ Absorptioncoefficient 0.764 mm⁻¹ F(000) 1056 Crystal prism; colorless Crystal size0.250 × 0.150 × 0.020 mm³ Theta range for data collection 3.769-62.794°Index ranges −9 ≦ h ≦ 9, −24 ≦ k ≦ 24, −15 ≦ l ≦ 16 Reflectionscollected 12795 Independent reflections 7704 [R_(int) = 0.0441]Absorption correction Semi-empirical from equivalents Max. and min.transmission 1.00000 and 0.80527 Refinement method Full-matrixleast-squares on F² Data/restraints/parameters 7704/52/705Goodness-of-fit on F² 1.048 Final R indices [F² > 2σ(F²)] R1 = 0.0658,wR2 = 0.1685 R indices (all data) R1 = 0.0805, wR2 = 0.1883 Absolutestructure parameter  −0.1 (2) Extinction coefficient 0.0030 (6) Largestdiff. peak and hole 0.327 and −0.228 e Å³

The asymmetric unit contains two molecules, denoted A and B, and fourwater positions, corresponding to a dihydrate. One water position issplit. The compound is a dihydrate. The absolute configuration at bothC9 and C9′ was established as the (5)-configuration byanomalous-dispersion effects. The Flack x parameters, −0.1(3), wasdetermined using 2360 quotients [(I+)−(I−)]/[(I+)+(I−)], as described inParsons, et al., Acta Cryst. 2013, B69, 249-259).

The crystal structure does not contain any apparent solvent-accessiblevoids. The crystal structure contains classical N—H • • • O, N—H • • • Nand O—H • • • O bonds (listed in Table 31).

TABLE 31 Hydrogen bonds in the Form III crystal structure. DHA angle D—H. . . A d(D—H) (Å) d(H . . . A) (Å) d(D . . . A) (Å) (°) N(19)—H(19A) .. . N(15′)#1 0.867 (15) 2.17 (2) 3.030 (7) 171 (7) N(25)—H(25) . . .O(2W)#2 0.861 (15) 2.16 (3) 2.964 (9) 155 (5) N(19′)—H(19C) . . .N(15)#3 0.866 (15) 2.214 (17) 3.079 (7) 176 (7) N(25′)—H(25′) . . .O(1W) 0.861 (15) 2.07 (3)  2.863 (10) 152 (5) O(1W)—H(1W) . . . O(34)0.84 (3) 1.92 (6) 2.714 (8)  157 (12) O(2W)—H(3W) . . . N(27)#4 0.81 (3)2.23 (7)  2.90 (4) 141 (8) O(2W)—H(4W) . . . O(34′) 0.82 (3) 2.13 (6)2.867 (9)  149 (10) O(3W)—H(5W) . . . O(35) 0.85 (3) 2.24 (7)  2.977(11)  144 (11) O(3W)—H(6W) . . . N(11′)#5 0.83 (3) 2.28 (7) 3.049 (9) 153 (13)Symmetry transformations used to generate equivalent atoms:#1 −x+1,y−1/2,−z+2 #2 x,y,z−1 #3 −x+1,y+1/2,−z+2 #4 x,y,z+1 #5−x+2,y−1/2,−z+1

Additional results, including atomic coordinates for the Form IIIcrystal structure, are given in Table 32, Table 33, and Table 34.

TABLE 32 Atomic coordinates (×10⁴), equivalent isotropic displacementparameters (Å² × 10³) and site occupancy factors for heavy atoms in theForm III crystal structure. U_(eq) is defined as one third of the traceof the orthogonalized U_(ij) tensor. Atom x y z Ueq s.o.f. C(1)  7254(13) 5083 (4) 7815 (5) 109 (3) 1 C(2) 6777 (9) 5436 (3) 6954 (5)  80 (2)1 C(3) 6374 (9) 5707 (3) 6235 (5)  78 (2) 1 C(4) 5962 (8) 6018 (3) 5343(4)  69 (2) 1 N(5) 4572 (6) 5852 (2) 4896 (4)  75 (1) 1 C(6)  4022 (10)6145 (4) 3994 (6) 105 (3) 1 C(7)  2525 (15) 5791 (7)  3682 (10) 121 (5)0.699 (17) C(8)  1854 (14)  5636 (13)  4614 (14) 128 (4) 0.699 (17)C(7A)  2244 (18)  6201 (15)  4070 (20) 121 (5) 0.301 (17) C(8A)  1860(30)  5600 (30)  4600 (40) 128 (4) 0.301 (17) C(9) 3334 (8) 5457 (3)5279 (6)  87 (2) 1 C(10) 3735 (8) 4767 (3) 5295 (5)  74 (2) 1 N(11) 4057(6) 4430 (2) 4554 (4)  71 (1) 1 C(12) 4209 (7) 3815 (3) 4838 (4)  63 (1)1 C(13) 3912 (7) 3769 (3) 5788 (4)  63 (1) 1 C(14) 3665 (7) 3278 (3)6463 (4)  65 (1) 1 N(15) 3411 (8) 3409 (3) 7350 (4)  82 (2) 1 C(16) 3324 (11) 4037 (4) 7594 (5)  96 (2) 1 C(17) 3371 (9) 4523 (3) 7006 (5) 87 (2) 1 N(18) 3645 (6) 4390 (2) 6065 (4)  69 (1) 1 N(19) 3695 (7) 2658(2) 6211 (4)  72 (1) 1 C(20) 4578 (7) 3318 (3) 4162 (4)  63 (1) 1 C(21)3901 (8) 3338 (3) 3233 (4)  75 (2) 1 C(22) 4268 (8) 2899 (3) 2575 (5) 82 (2) 1 C(23) 5353 (7) 2411 (3) 2823 (4)  70 (2) 1 C(24) 5740 (8) 1903(4) 2146 (5)  78 (2) 1 N(25) 5343 (7) 2023 (3) 1207 (4)  82 (2) 1 C(26)5402 (6) 1603 (2)  438 (3)  78 (2) 1 N(27) 5424 (6) 1895 (2) −399 (3) 90 (2) 1 C(28) 5407 (7) 1526 (3) −1180 (3)  108 (3) 1 C(29) 5320 (7) 880 (3) −1147 (4)  114 (3) 1 C(30) 5290 (7)  590 (2) −268 (5) 116 (3) 1C(31) 5335 (7)  952 (2)  556 (4) 104 (2) 1 C(32) 6035 (8) 2391 (3) 3738(4)  72 (2) 1 C(33) 5674 (8) 2835 (3) 4389 (4)  71 (2) 1 O(34) 6839 (6)6408 (2) 4997 (3)  92 (1) 1 O(35) 6352 (7) 1410 (3) 2427 (4) 105 (2) 1C(1′)  7696 (16) 2764 (5) 7523 (8) 142 (4) 1 C(2′)  7752 (12) 3245 (4)8271 (6) 104 (2) 1 C(3′)  7804 (11) 3640 (3) 8876 (6)  97 (2) 1 C(4′) 7786 (12) 4089 (3) 9650 (5)  87 (2) 1 N(5′) 9178 (7) 4331 (2) 9967 (4) 80 (2) 1 C(6′) 10718 (14)  4074 (10)  9711 (13) 109 (6) 0.614 (13)C(7′) 11884 (14) 4463 (6) 10289 (14) 125 (6) 0.614 (13) C(8′) 11090 (20) 4550 (20) 11219 (15) 108 (3) 0.614 (13) C(6″) 10790 (20)  4240 (20) 9650 (20) 109 (6) 0.386 (13) C(7″) 11720 (20)  4083 (10) 10698 (15) 125(6) 0.386 (13) C(8″) 10980 (40)  4550 (30) 11330 (30) 108 (3) 0.386 (13)C(9′) 9334 (7) 4700 (3) 10847 (4)   76 (2) 1 C(10′) 9125 (7) 5389 (3)10673 (4)   64 (1) 1 N(11′) 9385 (6) 5712 (2) 9891 (3)  63 (1) 1 C(12′)9236 (7) 6337 (3) 10105 (4)   58 (1) 1 C(13′) 8844 (6) 6404 (3) 11035(4)   60 (1) 1 C(14′) 8396 (7) 6902 (3) 11662 (4)   65 (1) 1 N(15′) 8097(6) 6786 (3) 12546 (3)   73 (1) 1 C(16′) 8185 (8) 6172 (3) 12851 (5)  77 (2) 1 C(17′) 8477 (8) 5671 (3) 12314 (4)   71 (2) 1 N(18′) 8795 (5)5792 (2) 11387 (3)   60 (1) 1 N(19′) 8247 (7) 7499 (2) 11341 (4)   74(1) 1 C(20′) 9595 (7) 6820 (2) 9394 (4)  58 (1) 1 C(21′) 8991 (8) 6766(3) 8469 (4)  74 (2) 1 C(22′) 9397 (8) 7207 (3) 7784 (4)  75 (2) 1C(23′) 10437 (7)  7698 (2) 8029 (4)  63 (1) 1 C(24′) 10911 (9)  8199 (3)7361 (5)  76 (2) 1 N(25′) 10449 (7)  8119 (3) 6439 (4)  76 (1) 1 C(26′)10663 (6)  8542 (2) 5682 (3)  90 (2) 1 N(27′) 10045 (6)  8313 (3) 4855(4) 114 (2) 1 C(28′) 10118 (8)  8689 (4) 4087 (3) 157 (6) 1 C(29′) 10828(9)  9274 (4) 4120 (5) 164 (6) 1 C(30′) 11467 (9)  9497 (2) 4989 (6) 147(5) 1 C(31′) 11393 (8)  9130 (3) 5801 (4) 112 (3) 1 C(32′) 11044 (8) 7747 (3) 8964 (4)  69 (2) 1 C(33′) 10637 (7)  7311 (3) 9641 (4)  66 (1)1 O(34′) 6502 (7) 4225 (2) 9989 (4) 100 (2) 1 O(35′) 11686 (9)  8659 (2)7651 (4) 119 (2) 1 O(1W) 9679 (9) 7013 (4) 5330 (5) 135 (2) 1 O(2W) 4409(8) 3159 (3) 10069 (4)  106 (2) 1 O(3W)  8713 (12)  472 (4) 1824 (8) 157(3) 1 O(4W) 14547 (14) 9678 (5) 7332 (8) 170 (5) 0.848 (14) O(4W′) 14510(80)  9400 (30)  8260 (50) 170 (5) 0.152 (14)

TABLE 33 Bond lengths (Å) and angles (°) for the Form III crystalstructure. C(1)—C(2)  1.456 (10) C(1)—H(1A) 0.96 C(1)—H(1B) 0.96C(1)—H(1C) 0.96 C(2)—C(3) 1.193 (9) C(3)—C(4) 1.438 (9) C(4)—O(34) 1.234(7) C(4)—N(5) 1.328 (8) N(5)—C(6) 1.458 (9) N(5)—C(9) 1.472 (8)C(6)—C(7)  1.499 (11) C(6)—C(7A)  1.509 (14) C(6)—H(6A) 0.97 C(6)—H(6B)0.97 C(6)—H(6A1) 0.97 C(6)—H(6A2) 0.97 C(7)—C(8)  1.510 (14) C(7)—H(7A)0.97 C(7)—H(7B) 0.97 C(8)—C(9)  1.544 (11) C(8)—H(8A) 0.97 C(8)—H(8B)0.97 C(7A)—C(8A)  1.523 (15) C(7A)—H(7A1) 0.97 C(7A)—H(7A2) 0.97C(8A)—C(9)  1.538 (14) C(8A)—H(8A1) 0.97 C(8A)—H(8A2) 0.97 C(9)—C(10)1.497 (9) C(9)—H(9) 0.98 C(9)—H(9A) 0.98 C(10)—N(11) 1.313 (8)C(10)—N(18) 1.357 (8) N(11)—C(12) 1.367 (7) C(12)—C(13) 1.389 (8)C(12)—C(20) 1.471 (8) C(13)—N(18) 1.396 (8) C(13)—C(14) 1.436 (8)C(14)—N(15) 1.317 (8) C(14)—N(19) 1.362 (8) N(15)—C(16)  1.378 (10)C(16)—C(17)  1.324 (11) C(16)—H(16) 0.93 C(17)—N(18) 1.396 (9)C(17)—H(17) 0.93 N(19)—H(19A)  0.867 (15) N(19)—H(19B)  0.864 (15)C(20)—C(21) 1.388 (9) C(20)—C(33) 1.395 (9) C(21)—C(22) 1.367 (9)C(21)—H(21) 0.93 C(22)—C(23)  1.404 (10) C(22)—H(22) 0.93 C(23)—C(32)1.372 (9) C(23)—C(24) 1.493 (9) C(24)—O(35) 1.216 (9) C(24)—N(25) 1.365(9) N(25)—C(26) 1.409 (6) N(25)—H(25)  0.861 (15) C(26)—N(27) 1.3363C(26)—C(31) 1.3898 N(27)—C(28) 1.3505 C(28)—C(29) 1.3707 C(28)—H(28)0.93 C(29)—C(30) 1.3874 C(29)—H(29) 0.93 C(30)—C(31) 1.3907 C(30)—H(30)0.93 C(31)—H(31) 0.93 C(32)—C(33) 1.366 (9) C(32)—H(32) 0.93 C(33)—H(33)0.93 C(1′)—C(2′)  1.468 (12) C(1′)—H(1′1) 0.96 C(1′)—H(1′2) 0.96C(1′)—H(1′3) 0.96 C(2′)—C(3′)  1.193 (10) C(3′)—C(4′)  1.450 (11)C(4′)—O(34′)  1.248 (10) C(4′)—N(5′)  1.321 (10) N(5′)—C(9′) 1.466 (7)N(5′)—C(6″)  1.470 (14) N(5′)—C(6′)  1.474 (12) C(6′)—C(7′)  1.475 (13)C(6′)—H(6′1) 0.97 C(6′)—H(6′2) 0.97 C(7′)—C(8′)  1.533 (14) C(7′)—H(7′1)0.97 C(7′)—H(7′2) 0.97 C(8′)—C(9′)  1.555 (11) C(8′)—H(8′1) 0.97C(8′)—H(8′2) 0.97 C(6″)—C(7″)  1.65 (5) C(6″)—H(6″1) 0.97 C(6″)—H(6″2)0.97 C(7″)—C(8″)  1.504 (14) C(7″)—H(7″1) 0.97 C(7″)—H(7″2) 0.97C(8″)—C(9′)  1.523 (13) C(8″)—H(8″1) 0.97 C(8″)—H(8″2) 0.97 C(9′)—C(10′) 1.488 (8) C(9′)—H(9′) 0.98 C(9′)—H(9″) 0.98 C(10′)—N(11′) 1.332 (8)C(10′)—N(18′) 1.366 (7) N(11′)—C(12′) 1.365 (7) C(12′)—C(13′) 1.388 (8)C(12′)—C(20′) 1.482 (8) C(13′)—N(18′) 1.389 (7) C(13′)—C(14′) 1.446 (8)C(14′)—N(15′) 1.316 (8) C(14′)—N(19′) 1.344 (8) N(15′)—C(16′) 1.370 (9)C(16′)—C(17′) 1.337 (9) C(16′)—H(16′) 0.93 C(17′)—N(18′) 1.383 (7)C(17′)—H(17′) 0.93 N(19′)—H(19C)  0.866 (15) N(19′)—H(19D)  0.862 (15)C(20′)—C(21′) 1.368 (8) C(20′)—C(33′) 1.385 (8) C(21′)—C(22′) 1.406 (9)C(21′)—H(21′) 0.93 C(22′)—C(23′) 1.384 (9) C(22′)—H(22′) 0.93C(23′)—C(32′) 1.381 (8) C(23′)—C(24′) 1.495 (8) C(24′)—O(35′) 1.223 (8)C(24′)—N(25′) 1.339 (9) N(25′)—C(26′) 1.417 (6) N(25′)—H(25′)  0.861(15) C(26′)—N(27′) 1.3318 C(26′)—C(31′) 1.3918 N(27′)—C(28′) 1.3508C(28′)—C(29′) 1.3735 C(28′)—H(28′) 0.93 C(29′)—C(30′) 1.3825C(29′)—H(29′) 0.93 C(30′)—C(31′) 1.3914 C(30′)—H(30′) 0.93 C(31′)—H(31′)0.93 C(32′)—C(33′) 1.392 (8) C(32′)—H(32′) 0.93 C(33′)—H(33′) 0.93O(1W)—H(1W)  0.84 (3) O(1W)—H(2W)  0.88 (3) O(2W)—H(3W)  0.81 (3)O(2W)—H(4W)  0.82 (3) O(3W)—H(5W)  0.85 (3) O(3W)—H(6W)  0.83 (3)C(2)—C(1)—H(1A) 109.5 C(2)—C(1)—H(1B) 109.5 H(1A)—C(1)—H(1B) 109.5C(2)—C(1)—H(1C) 109.5 H(1A)—C(1)—H(1C) 109.5 H(1B)—C(1)—H(1C) 109.5C(3)—C(2)—C(1) 177.9 (7) C(2)—C(3)—C(4) 176.8 (7) O(34)—C(4)—N(5) 121.1(6) O(34)—C(4)—C(3) 122.9 (6) N(5)—C(4)—C(3) 115.9 (5) C(4)—N(5)—C(6)120.5 (5) C(4)—N(5)—C(9) 126.4 (5) C(6)—N(5)—C(9) 112.3 (5)N(5)—C(6)—C(7) 104.1 (7) N(5)—C(6)—C(7A)  102.8 (10) N(5)—C(6)—H(6A)110.9 C(7)—C(6)—H(6A) 110.9 N(5)—C(6)—H(6B) 110.9 C(7)—C(6)—H(6B) 110.9H(6A)—C(6)—H(6B) 109 N(5)—C(6)—H(6A1) 111.2 C(7A)—C(6)—H(6A1) 111.2N(5)—C(6)—H(6A2) 111.2 C(7A)—C(6)—H(6A2) 111.2 H(6A1)—C(6)—H(6A2) 109.1C(6)—C(7)—C(8)  102.4 (10) C(6)—C(7)—H(7A) 111.3 C(8)—C(7)—H(7A) 111.3C(6)—C(7)—H(7B) 111.3 C(8)—C(7)—H(7B) 111.3 H(7A)—C(7)—H(7B) 109.2C(7)—C(8)—C(9)  104.3 (10) C(7)—C(8)—H(8A) 110.9 C(9)—C(8)—H(8A) 110.9C(7)—C(8)—H(8B) 110.9 C(9)—C(8)—H(8B) 110.9 H(8A)—C(8)—H(8B) 108.9C(6)—C(7A)—C(8A)  102.5 (15) C(6)—C(7A)—H(7A1) 111.3 C(8A)—C(7A)—H(7A1)111.3 C(6)—C(7A)—H(7A2) 111.3 C(8A)—C(7A)—H(7A2) 111.3H(7A1)—C(7A)—H(7A2) 109.2 C(7A)—C(8A)—C(9)  106.5 (14)C(7A)—C(8A)—H(8A1) 110.4 C(9)—C(8A)—H(8A1) 110.4 C(7A)—C(8A)—H(8A2)110.4 C(9)—C(8A)—H(8A2) 110.4 H(8A1)—C(8A)—H(8A2) 108.6 N(5)—C(9)—C(10)113.3 (6) N(5)—C(9)—C(8A) 102.2 (9) C(10)—C(9)—C(8A)   112 (2)N(5)—C(9)—C(8) 101.2 (7) C(10)—C(9)—C(8)  114.7 (13) N(5)—C(9)—H(9)109.1 C(10)—C(9)—H(9) 109.1 C(8)—C(9)—H(9) 109.1 N(5)—C(9)—H(9A) 109.8C(10)—C(9)—H(9A) 109.8 C(8A)—C(9)—H(9A) 109.8 N(11)—C(10)—N(18) 110.4(5) N(11)—C(10)—C(9) 125.3 (6) N(18)—C(10)—C(9) 123.9 (6)C(10)—N(11)—C(12) 107.7 (5) N(11)—C(12)—C(13) 109.3 (5)N(11)—C(12)—C(20) 120.7 (5) C(13)—C(12)—C(20) 129.9 (5)C(12)—C(13)—N(18) 104.5 (5) C(12)—C(13)—C(14) 137.7 (5)N(18)—C(13)—C(14) 117.4 (5) N(15)—C(14)—N(19) 117.3 (5)N(15)—C(14)—C(13) 121.5 (5) N(19)—C(14)—C(13) 121.1 (5)C(14)—N(15)—C(16) 117.2 (6) C(17)—C(16)—N(15) 126.0 (6)C(17)—C(16)—H(16) 117 N(15)—C(16)—H(16) 117 C(16)—C(17)—N(18) 117.1 (6)C(16)—C(17)—H(17) 121.4 N(18)—C(17)—H(17) 121.4 C(10)—N(18)—C(17) 132.0(5) C(10)—N(18)—C(13) 108.0 (5) C(17)—N(18)—C(13) 120.0 (5)C(14)—N(19)—H(19A)   118 (4) C(14)—N(19)—H(19B)   113 (4)H(19A)—N(19)—H(19B)   121 (3) C(21)—C(20)—C(33) 116.9 (5)C(21)—C(20)—C(12) 120.0 (6) C(33)—C(20)—C(12) 123.0 (5)C(22)—C(21)—C(20) 121.6 (6) C(22)—C(21)—H(21) 119.2 C(20)—C(21)—H(21)119.2 C(21)—C(22)—C(23) 120.5 (6) C(21)—C(22)—H(22) 119.7C(23)—C(22)—H(22) 119.7 C(32)—C(23)—C(22) 118.1 (6) C(32)—C(23)—C(24)119.1 (6) C(22)—C(23)—C(24) 122.7 (6) O(35)—C(24)—N(25) 122.8 (7)O(35)—C(24)—C(23) 121.1 (7) N(25)—C(24)—C(23) 116.1 (7)C(24)—N(25)—C(26) 127.6 (6) C(24)—N(25)—H(25)   119 (4)C(26)—N(25)—H(25)   112 (4) N(27)—C(26)—C(31) 124.7 N(27)—C(26)—N(25)113.2 (4) C(31)—C(26)—N(25) 121.9 (4) C(26)—N(27)—C(28) 117N(27)—C(28)—C(29) 123.1 N(27)—C(28)—H(28) 118.4 C(29)—C(28)—H(28) 118.4C(28)—C(29)—C(30) 118.4 C(28)—C(29)—H(29) 120.8 C(30)—C(29)—H(29) 120.8C(29)—C(30)—C(31) 120.3 C(29)—C(30)—H(30) 119.8 C(31)—C(30)—H(30) 119.8C(26)—C(31)—C(30) 116.3 C(26)—C(31)—H(31) 121.8 C(30)—C(31)—H(31) 121.8C(33)—C(32)—C(23) 121.0 (6) C(33)—C(32)—H(32) 119.5 C(23)—C(32)—H(32)119.5 C(32)—C(33)—C(20) 121.8 (6) C(32)—C(33)—H(33) 119.1C(20)—C(33)—H(33) 119.1 C(2′)—C(1′)—H(1′1) 109.5 C(2′)—C(1′)—H(1′2)109.5 H(1′1)—C(1′)—H(1′2) 109.5 C(2′)—C(1′)—H(1′3) 109.5H(1′1)—C(1′)—H(1′3) 109.5 H(1′2)—C(1′)—H(1′3) 109.5 C(3′)—C(2′)—C(1′) 179.5 (12) C(2′)—C(3′)—C(4′)  175.6 (10) O(34′)—C(4′)—N(5′) 123.3 (6)O(34′)—C(4′)—C(3′) 120.2 (8) N(5′)—C(4′)—C(3′) 116.6 (8)C(4′)—N(5′)—C(9′) 120.7 (6) C(4′)—N(5′)—C(6″)  131.2 (14)C(9′)—N(5′)—C(6″)  107.6 (13) C(4′)—N(5′)—C(6′) 122.9 (8)C(9′)—N(5′)—C(6′) 112.6 (7) N(5′)—C(6′)—C(7′)  102.4 (10)N(5′)—C(6′)—H(6′1) 111.3 C(7′)—C(6′)—H(6′1) 111.3 N(5′)—C(6′)—H(6′2)111.3 C(7′)—C(6′)—H(6′2) 111.3 H(6′1)—C(6′)—H(6′2) 109.2C(6′)—C(7′)—C(8′)  103.1 (13) C(6′)—C(7′)—H(7′1) 111.1C(8′)—C(7′)—H(7′1) 111.1 C(6′)—C(7′)—H(7′2) 111.1 C(8′)—C(7′)—H(7′2)111.1 H(7′1)—C(7′)—H(7′2) 109.1 C(7′)—C(8′)—C(9′)  101.6 (10)C(7′)—C(8′)—H(8′1) 111.4 C(9′)—C(8′)—H(8′1) 111.4 C(7′)—C(8′)—H(8′2)111.4 C(9′)—C(8′)—H(8′2) 111.4 H(8′1)—C(8′)—H(8′2) 109.3N(5′)—C(6″)—C(7″)  98.1 (18) N(5′)—C(6″)—H(6″1) 112.2 C(7″)—C(6″)—H(6″1)112.2 N(5′)—C(6″)—H(6″2) 112.2 C(7″)—C(6″)—H(6″2) 112.2H(6″1)—C(6″)—H(6″2) 109.8 C(8″)—C(7″)—C(6″)  102.1 (19)C(8″)—C(7″)—H(7″1) 111.3 C(6″)—C(7″)—H(7″1) 111.3 C(8″)—C(7″)—H(7″2)111.3 C(6″)—C(7″)—H(7″2) 111.3 H(7″1)—C(7″)—H(7″2) 109.2C(7″)—C(8″)—C(9′)  105.9 (12) C(7″)—C(8″)—H(8″1) 110.6C(9′)—C(8″)—H(8″1) 110.6 C(7″)—C(8″)—H(8″2) 110.6 C(9′)—C(8″)—H(8″2)110.6 H(8″1)—C(8″)—H(8″2) 108.7 N(5′)—C(9′)—C(10′) 112.4 (5)N(5′)—C(9′)—C(8″) 106.7 (8) C(10′)—C(9′)—C(8″)   112 (3)N(5′)—C(9′)—C(8′) 101.4 (8) C(10′)—C(9′)—C(8′)  110.5 (18)N(5′)—C(9′)—H(9′) 110.8 C(10′)—C(9′)—H(9′) 110.8 C(8′)—C(9′)—H(9′) 110.8N(5′)—C(9′)—H(9″) 108.7 C(10′)—C(9′)—H(9″) 108.7 C(8″)—C(9′)—H(9″) 108.7N(11′)—C(10′)—N(18′) 110.4 (4) N(11′)—C(10′)—C(9′) 128.0 (5)N(18′)—C(10′)—C(9′) 121.3 (5) C(10′)—N(11′)—C(12′) 106.8 (4)N(11′)—C(12′)—C(13′) 110.0 (5) N(11′)—C(12′)—C(20′) 119.5 (5)C(13′)—C(12′)—C(20′) 130.3 (5) C(12′)—C(13′)—N(18′) 105.0 (5)C(12′)—C(13′)—C(14′) 138.4 (5) N(18′)—C(13′)—C(14′) 116.4 (5)N(15′)—C(14′)—N(19′) 118.5 (5) N(15′)—C(14′)—C(13′) 121.5 (5)N(19′)—C(14′)—C(13′) 120.0 (5) C(14′)—N(15′)—C(16′) 117.9 (5)C(17′)—C(16′)—N(15′) 125.5 (6) C(17′)—C(16′)—H(16′) 117.2N(15′)—C(16′)—H(16′) 117.2 C(16′)—C(17′)—N(18′) 116.7 (6)C(16′)—C(17′)—H(17′) 121.6 N(18′)—C(17′)—H(17′) 121.6C(10′)—N(18′)—C(17′) 130.6 (5) C(10′)—N(18′)—C(13′) 107.8 (4)C(17′)—N(18′)—C(13′) 121.6 (5) C(14′)—N(19′)—H(19C)   117 (4)C(14′)—N(19′)—H(19D)   121 (4) H(19C)—N(19′)—H(19D)   120 (3)C(21′)—C(20′)—C(33′) 118.8 (5) C(21′)—C(20′)—C(12′) 120.7 (5)C(33′)—C(20′)—C(12′) 120.4 (5) C(20′)—C(21′)—C(22′) 120.7 (6)C(20′)—C(21′)—H(21′) 119.7 C(22′)—C(21′)—H(21′) 119.7C(23′)—C(22′)—C(21′) 120.6 (6) C(23′)—C(22′)—H(22′) 119.7C(21′)—C(22′)—H(22′) 119.7 C(32′)—C(23′)—C(22′) 118.2 (5)C(32′)—C(23′)—C(24′) 117.0 (5) C(22′)—C(23′)—C(24′) 124.7 (6)O(35′)—C(24′)—N(25′) 122.1 (6) O(35′)—C(24′)—C(23′) 120.9 (6)N(25′)—C(24′)—C(23′) 117.0 (6) C(24′)—N(25′)—C(26′) 127.6 (6)C(24′)—N(25′)—H(25′)   116 (3) C(26′)—N(25′)—H(25′)  116  (3)N(27′)—C(26′)—C(31′) 124.6 N(27′)—C(26′)—N(25′) 111.7 (4)C(31′)—C(26′)—N(25′) 123.7 (4) C(26′)—N(27′)—C(28′) 116.8N(27′)—C(28′)—C(29′) 123.4 N(27′)—C(28′)—H(28′) 118.3C(29′)—C(28′)—H(28′) 118.3 C(28′)—C(29′)—C(30′) 118.3C(28′)—C(29′)—H(29′) 120.8 C(30′)—C(29′)—H(29′) 120.8C(29′)—C(30′)—C(31′) 120.1 C(29′)—C(30′)—H(30′) 120 C(31′)—C(30′)—H(30′)120 C(30′)—C(31′)—C(26′) 116.7 C(30′)—C(31′)—H(31′) 121.7C(26′)—C(31′)—H(31′) 121.7 C(23′)—C(32′)—C(33′) 121.0 (5)C(23′)—C(32′)—H(32′) 119.5 C(33′)—C(32′)—H(32′) 119.5C(20′)—C(33′)—C(32′) 120.7 (5) C(20′)—C(33′)—H(33′) 119.7C(32′)—C(33′)—H(33′) 119.7 H(1W)—O(1W)—H(2W)   102 (4) H(3W)—O(2W)—H(4W)  116 (5) H(5W)—O(3W)—H(6W)   108 (5)

TABLE 34 Hydrogen coordinates [×10⁴] and isotropic displacementparameters [Å² × 10³] for the Form III crystal structure. Atom x y zU_(eq) s.o.f. H(1A) 7145 5347 8359 163 1 H(1B) 8348 4953 7804 163 1H(1C) 6585 4718 7850 163 1 H(6A) 4813 6101 3535 126 0.699 (17) H(6B)3799 6590 4079 126 0.699 (17) H(6A1) 4258 5882 3462 126 0.301 (17)H(6A2) 4508 6557 3927 126 0.301 (17) H(7A) 2762 5410 3337 145 0.699 (17)H(7B) 1792 6051 3286 145 0.699 (17) H(8A) 1109 5285 4542 153 0.699 (17)H(8B) 1310 5998 4858 153 0.699 (17) H(7A1) 1671 6215 3442 145 0.301 (17)H(7A2) 1984 6575 4420 145 0.301 (17) H(8A1) 1652 5254 4154 153 0.301(17) H(8A2) 925 5663 4946 153 0.301 (17) H(9) 3171 5596 5926 104 0.699(17) H(9A) 3144 5598 5921 104 0.301 (17) H(16) 3224 4129 8230 115 1H(17) 3227 4935 7212 105 1 H(19A) 3280 (80) 2380 (20) 6580 (30) 86 1H(19B) 3640 (80) 2600 (30) 5603 (12) 86 1 H(21) 3179 3658 3054 90 1H(22) 3795 2924 1958 99 1 H(25) 5230 (60) 2410 (11) 1030 (40) 50 (14) 1H(28) 5456 1720 −1768 129 1 H(29) 5282  642 −1701 137 1 H(30) 5239  153−229 139 1 H(31) 5321  768 1153 125 1 H(32) 6753 2070 3918 86 1 H(33)6172 2815 5000 85 1 H(1′1) 7861 2960 6926 213 1 H(1′2) 6672 2559 7482213 1 H(1′3) 8520 2456 7671 213 1 H(6′1) 10840 4124 9038 131 0.614 (13)H(6′2) 10824 3630 9877 131 0.614 (13) H(7′1) 12053 4866 9985 151 0.614(13) H(7′2) 12901 4247 10397 151 0.614 (13) H(8′1) 11562 4899 11589 1300.614 (13) H(8′2) 11152 4170 11600 130 0.614 (13) H(6″1) 11198 4619 9374131 0.386 (13) H(6″2) 10838 3890 9216 131 0.386 (13) H(7″1) 11527 365210891 151 0.386 (13) H(7″2) 12864 4150 10694 151 0.386 (13) H(8″1) 116224930 11400 130 0.386 (13) H(8″2) 10880 4370 11955 130 0.386 (13) H(9′)8585 4548 11294 91 0.614 (13) H(9″) 8518 4558 11258 91 0.386 (13) H(16′)8027 6096 13485 92 1 H(17′) 8467 5262 12553 85 1 H(19C) 7820 (80) 7770(20) 11710 (30)  89 1 H(19D) 8310 (90) 7590 (30) 10749 (16)  89 1 H(21′)8305 6435 8292 88 1 H(22′) 8963 7168 7160 90 1 H(25′) 10060 (50)  7754(12) 6280 (30) 44 (13) 1 H(28′) 9663 8544 3505 188 1 H(29′) 10879 95143572 197 1 H(30′) 11947 9893 5031 176 1 H(31′) 11809 9271 6393 135 1H(32′) 11735 8077 9144 83 1 H(33′) 11068 7350 10265 80 1 H(1W) 8700 (50)6930 (50) 5230 (80) 162 1 H(2W) 10010 (120) 6720 (40) 5740 (70) 162 1H(3W)  4760 (110) 2910 (30) 9700 (50) 128 1 H(4W)  4670 (120) 3529 (16)10000 (70)  128 1 H(5W)  8430 (150)  830 (30)  2010 (100) 189 1 H(6W) 9490 (110)  520 (60) 1500 (80) 189 1

Example 12 Comparison of Dissolution Rate and Exposure in Dogs for FreeBase Form I and Free Base Form II

The intrinsic dissolution rate (IDR) was measured for Forms I and II ofthe free base of Formula (1). DR was measured using a paddle overstationary disc equipped dissolution apparatus with concentrationdetermined using liquid chromatographic analysis against a standard. They-intercept normalized results are shown in FIG. 55 with the slopes andregression coefficient displayed. Form I has an IDR of 6.8 mg/cm²/min insimulated gastric fluid (SGF) (pH 1.2) and an IDR of 0.44 mg/cm²/min inpH 2.5 HCl/NaCl buffer. Form II has an DR of 5.4 mg/cm²/min in SGF andan DR of 0.35 mg/cm²/min in pH 2.5 HCl/NaCl buffer. Form I thereforeshows about a 26% increase in IDR at both conditions relative to FormII, which provides a significantly higher rate of dissolution that isadvantageous.

The plasma exposure of Forms I and II of the free base of Formula (1)were compared in nine fasted beagle dogs after a single oraladministration of 6 mg/kg of either form using batches with a similarparticle size distribution. The experiment was performed in 5 weeklyphases, with the Form II phase last. The area under the plasma drugconcentration-time curve (AUC), shown in FIG. 56, reflects the exposureto drug after administration of each preparation of Formula (1) and isexpressed in ng*h/L. Form II shows lower AUC than Form I in all dogs.Form II also shows lower C_(max) (maximum concentration) than Form I inall dogs. It was concluded that Form I had higher exposure in the beaglethan Form II. There was a good in vitro in vivo correlation for thedissolution rates of Form 1 and Form II, and the performance of eachform of Formula (1) when delivered by oral capsule to dogs. The superiorperformance of Form I in this dog study demonstrates that more favorabledosing is possible in humans relative to Form II.

Example 13 Screening for Cocrystals, Salts, and Mixed Salts/Cocrystalsof(S)-4-(8-Amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide

The cocrystal and salt formers given in Table 35 were employed inpreliminary screening experiments using a high-throughput approach.

TABLE 35 Summary of cocrystal and salt formers used in preliminaryscreening experiments. The codes for each salt/cocrystal former are usedin the batch identification numbers in the following sections.Salt/Co-crystal Former Code Formula g/mol Stock Solution Acetylsalicylicacid ASS C₉H₈O₄ 180.2 0.1M in acetone Adipic acid ADI C₆H₁₀O₄ 146.140.1M in acetone Capric acid CAP C₁₀H₂₀O₂ 172.27 0.1M in acetone Caprylicacid CPY C₈H₁₆O₂ 144.22 0.1M in acetone Gentisic acid GEN C₇H₆O₄ 154.120.1M in acetone Glutaric acid GLT C₅H₈O₄ 132.12 0.1M in acetone Glutaricacid, 2-Oxo OGL C₅H₆O₅ 146.1 0.1M in acetone Glycolic acid GLA C₂H₄O₃76.05 0.1M in acetone Lactic acid, L- LLA C₃H₆O₃ 90.08 0.1M in acetoneL-Malic acid MLA C₄H₆O₅ 134.09 0.1M in acetone Maionic acid MLO C₃H₄O₄104.06 0.1M in acetone Naphthoic acid, 1-hydroxy-2- XIN C₁₁H₈O₃ 188.180.1M in acetone Oxalic acid OXA C₂H₂O₄ 90.04 0.1M in acetoneToluenesulfonic acid TOS C₇H₈O₃S•H₂O 190.22 0.1M in acetonitrileCaffeine CAF C₈H₁₀N₄O₂ 194.19 0.1M in acetone:H₂O 9:1 Ethylmaltol ETMC₇H₈O₃ 140.14 0.1M in acetone Maltol MLL C₆H₆O₃ 126.11 0.1M in acetoneMenthol MEN C₁₀H₂₀O 156.27 0.1M in acetone Nicotinamide NCT C₆H₆N₂O122.13 0.1M in acetone Proline, L- PRO C₅H₉NO₂ 115.13 0.1M in methanol

The preliminary results are summarized in Table 36.

TABLE 36 Summary of the preliminary results of cocrystal and saltscreening experiments. Salt/Co-crystal Former Result Acetylsalicylicacid poor Adipic acid poor Capric acid poor Caprylic acid poor Gentisicacid excellent Glutaric acid poor Glutaric acid, 2-Oxo medium Glycolicacid poor Lactic acid, L- medium L-Malic acid medium Malonic acid goodNaphthoic acid, 1-hydroxy-2- good Oxalic acid excellent Toluenesulfonicacid medium Caffeine good Ethylmaltol medium Maltol medium Mentholmedium Nicotinamide poor Proline, L- very good Pyridoxine mediumSorbitol good Urea good Vanillin poor

More detailed screening experiments were performed. A total of 192high-throughput screening experiments were carried out in a microtiterplate (MTP) made from quartz. The list of salt and co-crystal formersused for the experimentation is given in Table 37, and extends beyondthe preliminary list in Table 35. The stock solutions of the 24 salt andco-crystal formers were filled into MTP well #A1-H3, A4-H6, A7-H9, andA10-H12. The same Formula (1) free base solution, which was prepared inTHF:water (9:1) at a concentration of 0.05 M, was filled into each ofthe 96 MTP wells.

TABLE 37 List of salt and co-crystal formers used as starting materialsin extended screening experiments. Salt/Co-crystal Former Code Formulag/mol Stock Solution Acetylsalicylic acid ASA C₉H₈O₄ 180.2 0.1M inacetone Adipic acid ADI C₆H₁₀O₄ 146.14 0.1M in acetone Capric acid CAPC₁₀H₁₂O₂ 172.27 0.1M in acetone Caprylic acid CPY C₈H₁₆O₂ 144.22 0.1M inacetone Gentisic acid GEN C₇H₆O₄ 154.12 0.1M in acetone Glutaric acidGLT C₅H₈O₄ 132.12 0.1M in acetone Glutaric acid, 2-oxo OGL C₅H₆O₅ 146.10.1M in acetone Glycolic acid GLA C₂H₄O₃ 76.05 0.1M in acetone Lacticacid, L- LLA C₃H₆O₃ 90.08 0.1M in acetone Malic acid, L- MLA C₄H₆O₅134.09 0.1M in acetone Malonic acid MLO C₃H₄O₄ 104.06 0.1M in acetoneNaphthoic acid, 1-hydroxy-2- XIN C₁₁H₈O₃ 188.18 0.1M in acetone Oxalicacid OXA C₂H₂O₄ 90.04 0.1M in acetone Toluenesulfonic acid TOSC₇H₈O₃S•H₂O 190.22 0.1M in acetonitrile Caffeine CAF C₈H₁₀N₄O₂ 194.190.1M in acetone:H₂O 9:1 Ethylmaltol ETM C₇H₈O₃ 140.14 0.1M in acetoneMaltol MLL C₆H₆O₃ 126.11 0.1M in acetone Menthol MEN C₁₀H₂₀O 156.27 0.1Min acetone Nicotinamide NCT C₆H₆N₂O 122.13 0.1M in acetone Proline, L-PRO C₅H₉NO₂ 115.13 0.1M in methanol Pyridoxine PYD C₈H₁₁NO₃ 169.18 0.1in isopropanol Sorbitol SBT C₆H₁₄O₆ 182.17 0.05M in methanol Urea URECH₄N₂O 60.06 0.1M in isopropanol Vanillin VLN C₈H₈O₃ 152.15 0.1M inacetone

The amount of substance per experiment was about 3 mg and the outcome ofthe crystallization experiments was evaluated by light microscopy. Sincethe stock solutions of the salt and co-crystal formers were generally0.1 M and the concentration of the stock solution of Form I of the freebase of Formula (1) was 0.05 M, 100 μL of free base stock solution wasmixed with 50 μL of salt or co-crystal former stock solution. The firstseries of 96 experiments the solvents were evaporated from the mixedstock solutions under a slight nitrogen flow at room temperature. Theresults of the microscopic investigation are summarized in Table 38.

TABLE 38 Results from visual (light microscopic) inspection of the MTPafter the first series of evaporation experiments. Abbreviations: amo =amorphous, pa-cry = partially crystalline, po-cry = possiblycrystalline. Salt/Co-crystal Former Well # Mic Well # Mic Well # MicWell # Mic Acetylsalicylic acid A1 amo A4 amo A7 amo A10 amo Adipic acidB1 amo B4 amo B7 amo B10 amo Capric acid C1 amo C4 amo C7 amo C10 amoCaprylic acid D1 amo D4 amo D7 amo D10 amo Gentisic acid E1 pa-cry E4pa-cry E7 cryst. E10 pa-cry Glutaric acid F1 amo F4 amo F7 amo F10 amoGlutaric acid, 2-oxo G1 amo G4 amo G7 amo G10 amo Glycolic acid H1 amoH4 amo H7 amo H10 amo Lactic acid, L- A2 amo A5 amo A8 amo A11 amo Malicacid, L- B2 amo B5 amo B8 amo B11 amo Malonic acid C2 amo C5 amo C8 amoC11 amo 1-hydroxy-2-naphthoic acid D2 amo D5 amo D8 amo D11 amo Oxalicacid E2 pa-cry E5 cryst E8 pa-cry E11 pa-cry Toluenesulfonic acid F2 amoF5 amo F8 amo F11 amo Caffeine G2 cryst G5 pa-cry G8 cryst G11 crystEthylmaltol H2 amo H5 amo H8 amo H11 amo Maltol A3 amo A6 amo A9 amo A12po-cry Menthol B3 amo B6 amo B9 amo B12 amo Nicotinamide C3 amo C6 amoC9 amo C12 amo Proline, L- D3 pa-cry D6 pa-cry D9 pa-cry D12 pa-cryPyridoxine E3 amo E6 amo E9 amo E12 amo Sorbitol F3 amo F6 pa-cry F9pa-cry F12 pa-cry Urea G3 pa-cry G6 pa-cry G9 pa-cry G12 pa-cry VanillinH3 amo H6 amo H9 amo H12 amo

After the first evaporation of the solvents the solids in the MTP weresubjected to a second series of 96 experiments, where four differentsolvent systems were explored. To the dry residues of the firstexperiment 100 μL of solvent according to Table 39.

TABLE 39 Solvents and solvent mixtures used for the second set ofexperiments. MTP well # Solvent System A1 through H3 acetone - water95:5 A4 through H6 isopropanol - water 95:5 A7 through H9 acetonitrileA10 through H12 ethyl acetate - water 99:1

The suspensions were agitated at 25° C. for two days then the solventswere evaporated under a slight nitrogen flow and the MTP was examined bylight microscopy. If the light microscopic image suggested the presenceof crystalline material, then a Raman spectrum was measured. Visuallyamorphous samples; e.g., glassy residues observed in the MTP were notfurther investigated. The results are given in Table 40.

TABLE 40 Results from light microscopy of the MTP after the second setof experiments. Abbreviations: amo = amorphous, pa-cry = partiallycrystalline, po-cry = possibly crystalline. Salt/Co-crystal Former Well# Mic Well # Mic Well # Mic Well # Mic Acetylsalicylic acid A1 amo A4amo A7 amo A10 amo Adipic acid B1 amo B4 amo B7 amo B10 amo Capric acidC1 amo C4 amo C7 amo C10 amo Caprylic acid D1 amo D4 amo D7 amo D10 amoGentisic acid E1 cryst E4 amo E7 cryst E10 cryst Glutaric acid F1 amo F4amo F7 amo F10 cryst Glutaric acid, 2-oxo G1 amo G4 amo G7 po-cry G10amo Glycolic acid H1 amo H4 amo H7 amo H10 amo Lactic acid, L- A2 po-cryA5 amo A8 amo A11 amo Malic acid, L- B2 amo B5 amo B8 amo B11 amoMalonic acid C2 po-cry C5 amo C8 po-cry C11 po-cry 1-hydroxy-2-naphthoicacid D2 po-cry D5 amo D8 po-cry D11 amo Oxalic acid E2 po-cry E5 amo E8cryst E11 pa-cry Toluenesulfonic acid F2 po-cry F5 amo F8 po-cry F11po-cry Caffeine G2 cryst G5 pa-cry G8 cryst G11 cryst Ethylmaltol H2 amoH5 amo H8 amo H11 cryst Maltol A3 amo A6 amo A9 amo A12 amo Menthol B3amo B6 amo B9 amo B12 amo Nicotinamide C3 amo C6 amo C9 amo C12 amoProline, L- D3 cryst D6 pa-cry D9 pa-cry D12 cryst Pyridoxine E3 po-cryE6 amo E9 amo E12 po-cry Sorbitol F3 amo F6 pa-cry F9 pa-cry F12 crystUrea G3 pa-cry G6 pa-cry G9 pa-cry G12 cryst Vanillin H3 amo H6 amo H9amo H12 amo

The result from the above experimentation is a rating of the lead foreach salt or co-crystal that was included in the screening program,which was classified as very good, good, intermediate, and poor. Saltand co-crystal leads that were classified as very good, good orintermediate are presented in Table 41. Poor leads were obtained foracetylsalicylic acid, adipic acid, capric acid, caprylic acid, glutaricacid, glycolic acid, nicotinamide, and vanillin; these are not listed inTable 41.

TABLE 41 Salt and co-crystal formers for which very good, good andintermediate leads were obtained. N is the number of scale-upexperiments (including additional suspension experiments). LeadSalt/Co-crystal former Code classification N Scale-up result Gentisicacid GEN very good 8 crystalline salt Glutaric acid, 2-oxo OGLintermediate 0 not applicable Lactic acid, L- LLA intermediate 0 notapplicable L-Malic acid MLA intermediate 0 not applicable Malonic acidMLO good 1 unsuccessful Naphthoic acid, 1-Hydroxy-2- XIN good 1amorphous Oxalic acid OXA very good 7 crystalline salt Toluenesulfonicacid TOS intermediate 0 not applicable Caffeine CAF good 1 unsuccessfulEthylmaltol ETM intermediate 0 not applicable Maltol MLL intermediate 0not applicable Menthol MEN intermediate 0 not applicable Proline, L- PROvery good 8 co-crystal Pyridoxine PYD intermediate 0 not applicableSorbitol SBT good 6 co-crystal Urea URE good 1 unsuccessful

Experiments with a significant number of potentially useful salt andco-crystal formers were unsuccessful. For example, experimentationproduced poor leads for acetylsalicylic acid, adipic acid, capric acid,caprylic acid, glutaric acid, glycolic acid, nicotinamide, and vanillin.Intermediate, good or very good leads can still turn out to be falseupon further examination, because crystallization in presence of othercompounds can potentially lead to a new crystal form or either compoundpresent in the experiment. Furthermore, amorphous solid residues wereobtained in all small scale experiments with adipic acid, ascorbic acid,aceturic acid (N-acetyl glycine), benzoic acid, nicotinic acid, andsaccharin.

Example 14 The L-Arabitol Cocrystal of(S)-4-(8-Amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideExample 14.1 Preparation of Form A of the L-Arabitol Cocrystal ofFormula (1)

The L-arabitol cocrystal of Formula (1) can be prepared by dissolvingboth components in methanol or in 96% ethanol at reflux temperature,followed by cooling the solution to room temperature. For instance, 1mmol of Formula (1) free base Form I and 2.25 mmol of L-arabitol aredissolved in methanol at reflux temperature and the solution is allowedto cool to room temperature. After stirring for several hours asuspension is obtained that contains the co-crystal (SP221-ARA-P3).

Example 14.2 Physical Characterization of Form A of the L-ArabitolCocrystal of Formula (1)

¹H NMR spectroscopy of the product from experiment SP221-ARA-P3 revealeda ratio of free base to L-arabitol of about 1:1.9 (or 1:2). Thisestimation is based on the sum of the integrals of the aromatic protonsof Formula (1) divided by the integral of two well separated protons ofL-arabitol at 4.2 and 4.3 ppm divided by 5.

Another sample of the L-arabitol cocrystal of Formula (1) (SP221-ARA-P4)was also examined by determination of CHNO content by elementalcomposition analysis. The molecular formula of a 1:2 co-crystal ofFormula (1) and L-arabitol would be C₃₆H₄₇N₇O₁₂ with a molecular weightof 769.8 g/mol. A dihydrate of this cocrystal would have the molecularformula of C₃₆H₅₁N₇O₁₄ and a molecular weight of 805.8 g/mol (with atheoretical water content of 4.5%). The results as presented in Table 42are in good agreement with the molecular formula for a 1:2 cocrystal (2moles of L-arabitol per mole of Formula (1)) dihydrate.

TABLE 42 Results from elemental composition analysis and water contentdetermination. Element % found Fit for C₃₆H₅₁N₇O₁₄ C 53.7 53.66 H 6.26.38 N 12.2 12.17 O 27.2 27.80 water 4.2 4.47

Based on elemental analysis and TG-FTIR mass loss, the stoichiometry ofthe co-crystal of Formula (1) with L-arabitol is a 1:2 cocrystaldihydrate (Form A) with the molecular formulaC₃₆H₅₁N₇O₁₄═[C₂₆H₂₃N₇O₂].2[C₅H₁₂O₅].2.H₂O and a molecular weight of805.8 g/mol.

The PXRD pattern of the product from experiment SP221-ARA-P3 isdisplayed in FIG. 57. The PXRD pattern of sample SP221-ARA-P3 wasverified against all known polymorphs of Formula (1) and the known PXRDpattern of L-arabitol. None of these forms can be detected in the PXRDpattern of the sample examined. All successfully produced samples of theco-crystal with L-arabitol exhibited the same PXRD pattern. Thefollowing peaks are characteristic of Form A of the 1:2 L-arabitolco-crystal of Formula (1): 5.6, 7.0, 9.2, 11.2, 12.9, 13.5, 14.0, 14.8,15.3, 16.8, 17.9, 18.2, 18.4, 21.1, 21.4, 22.5, 22.9, 23.9, 24.8, 25.5,26.0, 26.6, 27.7, 28.2, 28.8, and 29.4 °2θ±0.2 °2θ.

Optical microscopy of the L-arabitol cocrystal of Formula (1)(SP221-ARA-P4) shows crystalline material with predominantlyneedle-shaped particles with lengths of about 5 to 30 μm and widths ofabout 1 to 5 μm.

Raman spectroscopy was performed using a sample of the L-arabitolcocrystal of Formula (1) (SP221-ARA-P4). The Raman spectrum over themeasured range from 100 cm⁻¹ to 3500 cm⁻¹ and the fingerprint regionfrom 200 cm⁻¹ to 1800 cm⁻¹. The Raman spectrum was obtained in a similarmanner as described in Example 1.2 for Form I. Characteristic Ramanpeaks for Form A of the L-arabitol cocrystal of Formula (1) are observedat 2917, 2241, 1674, 1610, 1581, 1565, 1529, 1494, 1455, 1348, 1325,1309, 1264, 1242, 1189, 1164, 999, 872, and 279 (Raman shift, cm⁻¹±2cm⁻¹).

IR spectroscopy was also performed using a sample of the L-arabitolcocrystal of Formula (1) (SP221-ARA-P4). The IR spectrum over themeasured range from 600 cm⁻¹ to 3600 cm⁻¹ and the fingerprint regionfrom 600 cm⁻¹ to 1800 cm⁻¹. The IR spectrum was obtained in a similarmanner as described in Example 1.2 for Form I. Characteristic IR peaksfor Form A of the L-arabitol cocrystal of Formula (1) are observed at3471, 3188, 2924, 2239, 1670, 1637, 1621, 1603, 1579, 1524, 1505, 1435,1346, 1306, 1274, 1242, 1203, 1135, 1090, 1049, 1009, 998, 950, 902,892, 862, 821, 783, 739, 726, 711, 694, 637, and 621 (IR frequency,cm⁻¹±4 cm⁻¹).

The TG-FTIR thermogram suggests a water content of about 4.2%. This massloss is close to the water content for a dihydrate for which theexpected water content would be 4.47%. The DSC analysis in a closedsample pan reveals two endothermic events at about 110° C. and 127° C.(peak temperatures) that are followed by an exothermic signal with thepeak maximum at 157° C.

Dynamic vapor sorption analysis of the L-arabitol co-crystal (sampleSP221-ARA-P4) shows that the water content can strongly vary withchanging relative humidity. The water content at the end of the test isin good agreement with the TG-FTIR mass loss. The fact that all water isremoved at 0% RH suggests that the water is weakly bound in theco-crystal. The maximum water content of about 4.7% is reached at 95%RH. At the beginning of the test some excess water was present; however,the equilibrated water content at 50% RH is 4.1%. This result wascorroborated by a second DVS test of the same sample.

Example 15 The Citrate Salt of(S)-4-(8-Amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideExample 15.1 Preparation of Form A of Citrate Salt of Formula (1)

Citric acid has the molecular formula C₆H₈O₇ and a molecular mass of192.12 g/mol. The pK_(a) values of the three carboxylic acid groups incitric acid are 2.93, 4.76 and 6.40. Crystallization of the citrate saltfrom acetone-water mixtures led to samples that contained significantamounts of acetone and some water, while crystallization from 1-propanolled to a sample that contained a large amount of 1-propanol, indicatingthat both phases may be solvates.

Sample SP221-CIT-P4 was prepared as follows: 941 mg of free base ofFormula (1) (PP502-P1) and 384.5 mg citric acid was dissolved in 22 mLof acetone-water (10:1) by heating the mixture to 50° C. Upon cooling toroom temperature, a dilute suspension formed that was stirred in an openvial to let some solvent evaporate. More acetone was added, which led toa thicker suspension and which was filtered after stirring at roomtemperature for about one hour. About 436 mg of a white solid productwas obtained after drying in air at room temperature. The product fromexperiment SP221-CIT-P4 was further dried in air at 40° C. for 24 hoursto provide sample SP221-CIT-P4A. Batches produced by the procedures usedfor SP221-CIT-P4 and SP221-CIT-P4A may be additionally exposed tocontrolled humidity in order to exchange acetone for water.

Sample SP221-CIT-P6 was prepared as follows: 466 mg of free base ofFormula (1) (PP502-P1) and 96.4 mg citric acid was dissolved in 10 mL1-propanol 10:1 by heating the mixture to 70° C. At 50° C. the mixturewas seeded with SP221-CIT-P4 and let cool to room temperature. Asuspension formed from which the solid product was filtered off afterstirring at room temperature for about one hour. About 660 mg of a whitesolid product was obtained after drying in air at room temperature.Batches produced by the procedure used for SP221-CIT-P6 may beadditionally exposed to controlled humidity in order to exchange1-propanol for water.

Example 15.2 Physical Characterization of Form A of the Citrate Salt ofFormula (1)

¹H NMR spectroscopy of the product from experiment SP221-CIT-P4 revealeda ratio of Formula (1) to citric acid of about 2:1(1.83) based on thesum of the integrals for the 10 aromatic protons of Formula (1) dividedby the integral from the four protons from the methylene groups ofcitric acid between 2.6 and 2.9 ppm. The 1:2 citric acid:Formula (I) maybe a phase containing both ionized citric acid (as in a salt) andnon-ionized citric acid (as in a cocrystal). The molecular formula of a2:1 salt or co-crystal of Formula (1) with citric acid is2.[C₂₆H₂₃N₇O₂]+C₆H₈O₇ with a molecular weight of 1123.1 g/mol. In aninitial attempt to convert the acetone solvate into a hydrate sample,SP221-CIT-P4 was subjected to suspension equilibration in water at 25°C. for 24 hours, which resulted in conversion to Form III of the freebase of Formula (1) (the dihydrate).

Reflection PXRD patterns of the citrate salt obtained from acetone-water(SP221-CIT-P4) and 1-propanol (SP221-CIT-P6) are shown in FIG. 58. Anoverlay of the two PXRD patterns illustrates that the PXRD patterns ofthe two forms show remarkable similarities, indicating a similar crystallattice for both samples, and thus the two patterns likely represent twodifferent solvation states of a single host structure comprising citrateand Formula (1). Both samples are therefore designated Form A. Form A ofthe citrate salt of Formula (1) can also include other small organicsolvents and water in variable amounts. The following peaks arecharacteristic of Form A of the citrate salt of Formula (1), when in theapproximate solvation state of sample SP221-CIT-P4a: 6.1, 6.6, 7.2, 7.9,8.3, 9.7, 10.8, 11.1, 12.2, 13.5, 14.1, 14.9, 15.9, 16.6, 17.5, 17.9,18.3, 18.9, 19.5, 20.3, 21.5, 21.9, 22.7, 23.8, 24.4, 24.8, 26.1, 26.3,27.2, 27.4, 27.9, and 29.3 °2θ±0.2 °2θ. The following peaks arecharacteristic of Form A of the citrate salt of Formula (1), when in theapproximate solvation state of sample SP221-CIT-P6: 6.1, 6.4, 7.2, 7.9,8.2, 9.6, 10.9, 12.0, 13.4, 13.8, 14.0, 14.9, 15.5, 15.9, 16.4, 17.3,17.5, 18.2, 18.6, 19.3, 20.1, 20.4, 21.4, 21.6, 22.6, 23.2, 23.7, 24.3,26.0, 27.0, 27.3, 27.8, and 29.2 °2θ±0.2 °2θ. The foregoingcharacteristic peaks may vary in their position with the exchange ofsolvent into this crystalline phase.

Raman spectroscopy over a spectral range of 100 cm⁻¹ to 3500 cm⁻¹, ofForm A of the citrate salt of Formula (1) was performed on samplesSP221-CIT-P4 a Characteristic Raman peaks for Form A of the citrate saltof Formula (1), when in the approximate solvation state of sampleSP221-CIT-P4a, are observed at 3068, 2921, 2237, 1682, 1612, 1551, 1505,1436, 1332, 1313, 1241, 1188, 993 and 712 (Raman shift, cm⁻¹±2 cm⁻¹)(FIG. 59). Characteristic Raman peaks for Form A of the citrate salt ofFormula (1), when in the approximate solvation state of sampleSP221-CIT-P6, are observed at 3055, 2920, 2237, 1685, 1612, 1549, 1504,1436, 1333, 1313, 1286, 1240, 1187, 993 and 712 (Raman shift, cm⁻¹±2cm⁻¹). The foregoing characteristic peaks may vary in their positionswith the exchange of solvent into this crystalline phase.

ATR-IR spectroscopy of Form A, measured from 600 cm⁻¹ to 3600 cm⁻¹, ofthe citrate salt of Formula (1) was performed on samples SP221-CIT-P4aand SP221-CIT-P6. Characteristic IR peaks for Form A of the citrate saltof Formula (1), when in the approximate solvation state of sampleSP221-CIT-P4a, are observed at 3396, 2234, 1673, 1606, 1537, 1428, 1304,1264, 1200, 1092, 1008, 893, 866, 773, 735, and 693 (IR frequency,cm⁻¹±4 cm⁻¹). Characteristic IR peaks for Form A of the citrate salt ofFormula (1), when in the approximate solvation state of sampleSP221-CIT-P6, are observed at 3403, 2960, 2872, 2233, 1678, 1608, 1582,1538, 1434, 1403, 1352, 1302, 1253, 1201, 1094, 1055, 1010, 967, 895,813, 772, 750, 735, 693, and 612 (IR frequency, cm⁻¹±4 cm⁻¹). Theforegoing characteristic peaks may vary in their positions with theexchange of solvent into this crystalline phase.

TG-FTIR analysis was carried out on three different samples of thecitrate salt. The results show that the sample contains both water andacetone and that the water is less strongly bound than the acetone. Asample of SP221-CIT-P4 was stored for three months after preparation atambient conditions and denoted sample SP221-CIT-P3. TG-FTIR analysis ofthis sample revealed that most of the mass loss was due to water. Thisprovides evidence that acetone was slowly replaced by water over time,with a water content increase to about 8%. This observation is supportedby the finding that typically the mass loss occurs in two steps. In thefirst step water and some acetone is released and in the second step themass loss is predominantly due to acetone. The theoretical acetonecontent for a Formula (1):citrate 2:1 salt acetone monosolvate would be5% and the theoretical water content for a pentahydrate would be 8%.Therefore, in addition to an acetone solvate (or a mixed acetonesolvate-hydrate), a pure hydrated state of Form A of the citrate salt ofFormula (1) can be prepared. The result from TG-FTIR analysis of the1-propanol solvate shows two distinct steps, which may indicate that asecond 1-propanol solvate phase with a different stoichiometry exists.

Sample SP221-CIT-P3 of Form A of the citrate salt of Formula (1) wasselected for a DSC test in a closed sample pan and was observed toexhibit a broad endotherm that obscured melting. For a second DSCexperiment, the citrate sample SP221-CIT-P3 was stored under 33%relative humidity for several days of equilibration. The resulting DSCthermogram did not show any significant differences. The maximum of theendothermic signal is at 90° C.; however, the deviation from thebaseline begins even below 60° C. and a pronounced shoulder is found atabout 82° C. The exotherm that begins at about 140° C. is likely theresult of thermal degradation.

Dynamic vapor sorption (DVS) analysis of the citrate salt (sampleSP221-CIT-P4) shows that the given salt form absorbs a substantialamount of water at high humidity conditions and that at the end of thetest the water content is about 7.5% by weight. It is likely that partof the acetone that was found by TG-FTIR was exchanged by water duringthe DVS test.

Example 16 The Gentisate Salt of(S)-4-(8-Amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideExample 16.1 Preparation of Form A and Other Forms of the Gentisate Saltof Formula (1)

Gentisic acid has the chemical name 2,5-dihydroxybenzoic acid, themolecular formula C₇H₆O₄, and a molecular mass of 154.12 g/mol. ThepK_(a) of gentisic acid is 2.93. The gentisate salt was first identifiedin the screen described above (sample SP221-GEN-P1) and was reproducedby crystallization from an acetone-water mixture as an acetonehemisolvate (SP221-GEN-P2). Suspension equilibration of the acetonehemisolvate in acetonitrile led to a crystalline sample that did notcontain residual organic solvent (sample SP221-GEN-P3). TG-FTIR showedthat this sample contained about 2.6% of water. This result agreed wellwith the theoretical water content for a gentisate monohydrate of 2.8%.

Sample SP221-GEN-P1 was prepared as follows: 235.6 mg of Formula (1)free base (PP502-P1, 0.5 mmol) was dissolved in 4.0 mL of acetone-water(9:1) at 57° C. and 5.0 mL of a 0.1 M stock solution of gentisic acid inacetone was added. The mixture was allowed to cool to room temperatureand stirred while the cap was kept open to let acetone evaporate. Aftera suspension with a volume of about 3 mL was obtained, the solid productwas filtered off and dried in air at room temperature.

Sample SP221-GEN-P2 was prepared as follows: 470 mg of Formula (1) freebase (PP502-P1, 0.5 mmol) was dissolved in 11.0 mL of a 0.1 M stocksolution of gentisic acid in acetone. 2.0 mL of water was added to thissolution. The solution was seeded with a small amount of SP221-GEN-P1and stirred in an open vial to allow solvent evaporation. The solutionwas allowed to cool to room temperature and was stirred while the capwas left open to continue to allow acetone to evaporate. After asuspension with a volume of about 3 mL was obtained, the solid productwas filtered off and dried in air at room temperature.

Sample SP221-GEN-P3 was prepared as follows: 2.0 mL of acetonitrile wasadded to 58 mg of sample SP221-GEN-P2, and the resulting suspension wasstirred at room temperature for three days. The solids were filtered anddried in air at room temperature.

Sample SP221-GEN-P4 was prepared as follows: 466 mg of PP502-P1 (0.5mmol) and 154 mg of gentisic acid dissolved in 10.0 mL of 2-propanol byheating to 70° C. To facilitate dissolution, 0.2 mL of formic acid wasadded. The solution was allowed to cool to room temperature and wasseeded with SP221-GEN-P2 at about 45° C., and 5.0 mL of 2-propanol wasadded. Within about four hours a suspension was obtained from which thesolid product was filtered off and dried in air at room temperature.

An additional batch (sample SP221-GEN-P5) of the gentisate salt ofFormula (1) monohydrate was prepared by a similar method as was used toprepare SP221-GEN-P3. To about 400 mg of sample SP221-GEN-P4 was added4.2 mL of acetonitrile containing 5% water. The resulting suspension wasstirred at room temperature for one day. The solids were filtered offand dried product in air at room temperature. Sample SP221-GEN-P5A wasprepared by keeping sample SP221-GEN-P5 at 33% relative humidity for twoweeks.

A second hydrate, possibly a dihydrate, was obtained as the solidresidue after a solubility test (sample SP221-GEN-P6).

Example 16.2 Physical Characterization of Form A and Other Forms of theGentisate Salt of Formula (1)

¹H NMR spectroscopy of the product from experiment SP221-GEN-P5 revealeda ratio of Formula (1) free base to gentisic acid of 1:1 based upon thesum of integrated signals of four aromatic protons of Formula (1)between 7.5 and 8.5 ppm and two aromatic protons of gentisic acid thatappear between 6.6 and 7.0 ppm. The ¹H NMR spectrum also confirmed thatthe obtained material is essentially free of organic solvent.

In addition, sample SP221-GEN-P5 was analyzed for CHNO content byelemental composition analysis. The molecular formula of a 1:1 salt ofFormula (1) with gentisic acid is expected to be C₃₃H₂₉N₇O₆ with amolecular weight of 619.6 g/mol. A monohydrate of a 1:1 salt of Formula(1) with gentisic acid would have the molecular formula of C₃₃H₃₁N₇O₇and a molecular weight of 637.65 g/mol (with a water content of 2.8%).The results as presented in Table 43 are in agreement with the expectedformula for a monohydrate.

TABLE 43 Results from elemental composition analysis and water contentdetermination for sample SP221-GEN-P5. Element % found C₃₃H₃₁N₇O₇ C 60.962.16 H 5.1 4.90 N 15.0 15.38 O 16.5 17.56 water 2.6* 2.82 *This valuewas taken from the TG-FTIR analysis of sample SP221-GEN-P3, as describedbelow.

Optical microscopy of the gentisate salt of Formula (1) monohydrate(sample SP221-GEN-P5) showed crystalline material with predominantlyneedle-shaped particles with lengths of about 5 to 50 μm and widths ofabout 1 to 10 μm.

A reflection PXRD pattern of the gentisate salt of Formula (1)monohydrate is shown in FIG. 60 (sample SP221-GEN-P3). The PXRD patternof sample SP221-GEN-P5 (not shown) was indistinguishable from thepattern of sample SP221-GEN-P3, indicating that both samples arerepresentative of the same crystalline phase. This crystalline phase isdesignated Form A (monohydrate) of the gentisate salt of Formula (1).The following peaks are characteristic of Form A (monohydrate) of thegentisate salt of Formula (1): 4.6, 8.2, 9.0, 9.7, 11.8, 12.9, 13.8,14.5, 15.5, 16.6, 16.8, 18.4, 19.6, 20.5, 21.1, 24.1, 24.5, 25.5, 25.8,26.0, 26.6, 26.9, 27.4, and 29.8 °2θ±0.2 °2θ.

Raman spectroscopy was measured in a range from 100 cm⁻¹ to 3500 cm⁻¹using a sample of Form A (monohydrate) of the gentisate salt of Formula(1) (sample SP221-GEN-P5). The Raman spectrum was obtained in a similarmanner as described in Example 1.2 for Form I. Characteristic Ramanpeaks for Form A (monohydrate) of the gentisate salt of Formula (1) areobserved at 3057, 2919, 2223, 1681, 1613, 1576, 1552, 1518, 1437, 1333,1312, 1228, 1192, 1156, 990, 716, 485, and 257 (Raman shift, cm⁻¹±2cm⁻¹).

IR spectroscopy was measured in a range from 600 cm⁻¹ to 3600 cm⁻¹ usinga sample of Form A (monohydrate) of the gentisate salt of Formula (1)(sample SP221-GEN-P5). The IR spectrum was obtained in a similar manneras described in Example 1.2 for Form I. Characteristic IR peaks for FormA (monohydrate) of the gentisate salt of Formula (1) are observed at2957, 1682, 1668, 1602, 1574, 1523, 1504, 1481, 1429, 1377, 1346, 1302,1274, 1228, 1157, 1092, 1010, 939, 896, 865, 826, 810, 778, 748, 734,686, 660, and 617 (IR frequency, cm⁻¹±4 cm⁻¹).

The TG-FTIR thermogram of Form A (monohydrate) was measured for thegentisate salt of Formula (1) (sample SP221-GEN-P3). Differentialscanning calorimetry of Form A (monohydrate) of the gentisate salt ofFormula (1) (sample SP221-GEN-P5A), revealed two small endothermic peaksat 106° C. and 121° C. These peaks are unlikely to correspond to meltingof the salt, but can be assigned to phase transformations. The deviationfrom the baseline at 180° C. is tentatively attributed to the beginningof a melting process; however, thermal degradation is the dominatingphenomenon above 195° C. and a distinct melting point could not beidentified by DSC.

DVS analysis of Form A (monohydrate) of the gentisate salt of Formula(1) (sample SP221-GEN-P5) reveals several steps when scanning from highto lower RH. This suggests that more than one hydrate might exist. Eventhough the observed hysteresis is not symmetrical, a second test of thesame sample has shown that the entire DVS hydration-dehydration cycle isreversible. PXRD of the sample recovered from the DVS sample pan showedthe same pattern as the solid residue of the solubility experiments. Thewater content of 5.2% essentially corresponds to the water content of adihydrate. The following peaks are characteristic of the dihydrategentisate salt of Formula (1) obtained after the DVS test: 4.6, 8.7,11.7, 12.5, 12.8, 13.1, 14.1, 15.1, 15.6, 16.5, 16.8, 19.7, 24.1, 24.5,25.3, 25.7, 25.9, 26.6, 26.9, and 29.4 °2θ±0.2 °2θ.

Example 17 Characterization of the Oxalate Salt of(S)-4-(8-Amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideExample 17.1 Preparation of Form A of the Oxalate Salt of Formula (1)

Oxalic acid has the molecular formula C₂H₂O₄ with a molecular mass of90.04 g/mol. The pKa values of the two acid groups are 1.27 and 4.27.The oxalate salt was first identified in the screen described above.

Sample SP221-OXA-P1 was prepared as follows: 236 mg of sample PP502-P1and 45.4 mg of oxalic acid (Sigma Aldrich #75688) were added to 5.0 mLof acetone:water (95:5) and the mixture was heated to about 55° C. Thecompound did not dissolve; the mixture was allowed to cool to roomtemperature and stirred overnight, after which the solid was filteredoff and dried in air at room temperature.

Sample SP221-OXA-P2 was prepared as follows: 468.2 mg of sample PP502-P1and 90.9 mg of oxalic acid (Sigma Aldrich #75688) was added to 10.0 mLof 1-propanol and heated to 70° C. An essentially unstirable gel wasobtained to which another 15.0 mL of 1-propanol and 1.0 mL of water wereadded. Stirring was continued at room temperature for three days beforethe solid was filtered off and examined by PXRD after short drying inair at room temperature.

Sample SP221-OXA-P3 was prepared as follows: 468 mg of sample PP502-P1was dissolved in 10.0 mL of methanol at reflux, and 90 mg of oxalic aciddissolved in 2.0 mL of methanol was added. The material was cooled toroom temperature, seeded with SP221-OXA-P2, and about half of thesuspension was taken and stirred at room temperature then heated againto 50° C.; after which all solids dissolved. The solution was allowed tocool to room temperature again and stirred before part of the sample wasfiltered and solid investigated after drying in air at room temperature.This sample was designated SP221-OXA-P3A. To the other half of thesuspension from was added 3.0 mL of water. All solid immediatelydissolved, then the mixture was stirred under nitrogen purge at roomtemperature until all solvents were removed. To the dry residue wasadded 2.0 mL of acetonitrile, 2.0 mL of ethanol and 0.2 mL of water andstirred for two days at room temperature. A suspension was obtained fromwhich the solid was filtered off and dried in air at room temperature.This sample was designated SP221-OXA-P3B.

Sample SP221-OXA-P4 was prepared as follows: 468 mg of sample PP502-P1was dissolved in 10.0 mL of acetone and 1.0 mL of water at reflux, and45 mg of oxalic acid (Sigma Aldrich #75688) dissolved in 1.0 mL of waterwas added. No crystallization was observed. An additional 46 mg of solidoxalic acid and 5.0 mL of acetone were added and stirring at roomtemperature was continued while the vial was kept open. After overnightstirring, a thick paste was obtained. Heating the mixture to 50° C. ledto complete dissolution and cooling to room temperature again led to avery thick suspension. Part of the suspension was filtered and the soliddried in air at room temperature.

Sample SP221-OXA-P5 was prepared as follows: 470 mg of sample PP502-P1and 90 mg of oxalic acid (Sigma Aldrich #75688) were combined in 10.0 mLof tetrahydrofuran and 1.0 mL of methanol heated to reflux to achievedissolution of the solids. Upon seeding with SP221-OXA-P2 and cooling toroom temperature, a thick paste was obtained that was heated to 60° C.to 65° C. and stirred for two days before the solid was filtered off anddried in air at room temperature.

Sample SP221-OXA-P7 was prepared as follows: the remaining products fromexperiments SP221-OXA-P4 and SP221-OXA-P5 (about 300 mg) were combinedand were suspended in 5.0 mL of water. The mixture was stirred at roomtemperature for four days. The suspension was filtered and the solidswere dried in air at room temperature for 24 hours.

Example 17.2 Physical Characterization of Form A of the Oxalate Salt ofFormula (1)

¹H NMR spectroscopy of the oxalate was not carried out because of thelack of non-exchangeable hydrogens in oxalic acid. The CHNO content ofSP221-OXA-P1 was determined by elemental composition analysis, with theresults shown in Table 44. The molecular formula of a 1:1 salt ofFormula (1) with oxalic acid is predicted to be C₂₈H₂₅N₇O₆ with amolecular weight of 555.55 g/mol. A hydrate with a stoichiometry of 2.5moles of water to 1 mole of Formula (1) would have the molecular formulaof C₂₈H₃₀N₇O_(8.5) and a molecular weight of 602.6 g/mol (with a watercontent of 7.5%). The results as presented for sample SP221-OXA-P7 arein fair agreement with the molecular formula for such a “2.5 hydrate.”An assumed water content of about 7.5% is based on the result fromTG-FTIR which revealed a mass loss of 8.3% essentially attributable towater. A trihydrate would contain 8.9% of water and therefore atrihydrate is also possible.

TABLE 44 Results from elemental composition analysis of oxalate saltsand theoretical compositions. % found for % found for % theoretical %theoretical % theoretical SP221-OXA- SP221-OXA- for anhydrous for for2.5 Element P1 P7 form monohydrate hydrate C 57.4 53.8 60.54 58.63 55.81H 4.9 5.3  4.54  4.74  5.35 N 16.6 15.9 17.65 17.09 16.27 O 17.4 21.117.28 19.53 22.57 Sum 96.3% 96.1% 100% 100% 100%

A reflection PXRD pattern of the oxalate salt of Formula (1) monohydrateis shown in FIG. 61 (sample SP221-OXA-P7). This crystalline phase isdesignated Form A (2.5 hydrate) of the oxalate salt of Formula (1). Thefollowing peaks are characteristic of Form A (2.5 hydrate) of theoxalate salt of Formula (1): 5.5, 5.8, 7.4, 9.3, 11.0, 11.5, 12.7, 15.2,16.5, 17.3, 18.5, 18.7, 19.1, 19.7, 20.2, 20.8, 22.0, 22.3, 23.3, 23.6,24.8, 27.4, 28.6, 29.3, 29.6, 31.2, and 33.1 °2θ±0.2 °2θ.

Raman spectroscopy was measured over the range from 100 cm⁻¹ to 3500cm⁻¹ using a sample of Form A (2.5 hydrate) of the oxalate salt ofFormula (1) (sample SP221-OXA-P7). The Raman spectrum was obtained in asimilar manner as described in Example 1.2 for Form I. CharacteristicRaman peaks for Form A (2.5 hydrate) of the oxalate salt of Formula (1)are observed at 3073, 2992, 2950, 2922, 2247, 1671, 1612, 1584, 1552,1504, 1469, 1440, 1336, 1311, 1273, 1235, 1191, 1162, 1095, 1012, 897,718, 633, 409, 370, and 263 (Raman shift, cm⁻¹±2 cm⁻¹).

IR spectroscopy was measured over the range from 600 cm⁻¹ to 3600 cm⁻¹using a sample of Form A (2.5 hydrate) of the oxalate salt of Formula(1) (sample SP221-OXA-P7). The IR spectrum was obtained in a similarmanner as described in Example 1.2 for Form I. Characteristic IR peaksfor Form A (2.5 hydrate) of the oxalate salt of Formula (1) are observedat 3419, 2249, 1670, 1615, 1544, 1503, 1438, 1391, 1334, 1304, 1262,1195, 1151, 1126, 1093, 1013, 894, 877, 823, 783, 765, 738, and 652 (IRfrequency, cm⁻¹±4 cm⁻¹).

The TG-FTIR thermogram of Form A (2.5 hydrate) of the oxalate salt ofFormula (1) (sample SP221-OXA-P7) was measured. The observed mass loss,likely due to water, is between the expected water content for atrihydrate (8.9%) and a dihydrate (6.1%). The water appears to be weaklybound, because the beginning of the mass loss is essentially at roomtemperature. Differential scanning calorimetry results for Form A (2.5hydrate) of the oxalate salt of Formula (1) (sample SP221-OXA-P7). DSCrevealed a melting endotherm at 127° C. with an enthalpy of fusion ofabout 70 J/g. Thermal events observed in the DSC above 150° C. arelikely due to thermal decomposition.

DVS analysis of Form A (2.5 hydrate) of the oxalate salt of Formula (1)(sample SP221-OXA-P7) reveals that the water is removed at 0% RH. PXRDof the sample recovered from the DVS sample pan showed the same patternas pattern before the start of the DVS test. At 50% RH, the watercontent is about 5.5% and at 95% RH, the water content is about 6.5%.This result suggests that Form A may form a stable dihydrate.

Two additional PXRD patterns were obtained from preparations of oxalatesalts. PXRD patterns of samples SP221-OXA-P3B and SP221-OXA-P4 are shownin FIG. 62. Based on their PXRD patterns, these samples likely representother crystalline oxalate salts of Formula (1).

Example 18 Characterization of the L-Proline Cocrystal of(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideExample 18.1 Preparation of Form A of the L-Proline Cocrystal of Formula(1)

Cocrystals of Formula (1) with L-proline was prepared as follows. SampleSP221-PRO-P1 was prepared using 235 mg of Formula (1) free base(PP502-P1) and 57.5 mg L-proline, which were dissolved in 6.0 mL ofacetone-water (95:5), after which the solution was heated to about 58°C. An essentially clear solution was obtained, that was allowed to coolto room temperature and stirred overnight. After about 20 hours theobtained suspension was filtered and the solid product dried in air atroom temperature.

Sample SP221-PRO-P2 was prepared as follows: 41.2 mg of L-proline wasdissolved in 5.0 mL of ethanol containing about 1% of water. 168 mg ofFormula (1) free base (PP502-P1) was dissolved in 7.0 mL of ethanol atabout 60° C. The L-proline solution was then added to the Formula (1)solution, and the combined solution was allowed to cool to roomtemperature and stirred overnight. About 5 mL of ethanol was allowed toevaporate, and the solids were filtered.

Sample SP221-PRO-P3 was prepared as follows: 235 mg PP502-P1 and 80.4 mgof L-proline were suspended in 8.0 mL of ethanol and heated to 70° C.Another 1.5 ml of ethanol was added at 70° C. in order to obtain anessentially clear solution. The solution was allowed to cool to roomtemperature within about one hour and was stirred at room temperaturefor about 2 hours. The solids were filtered and the solid product wasdried in air at room temperature.

Sample SP221-PRO-P6 was prepared as follows: 940 mg of PP502-P1 wasdissolved in 15.0 mL of ethanol and 1.0 mL of water by heating to about75° C., and 345 mg of L-proline was dissolved in 4.0 mL ethanol:water(3:1) and combined. The solution was allowed to cool to 40° C. andstirred at 40° C. under a steady flow of nitrogen. Over three days allsolvents evaporated and a dry residue was obtained. The solid wasresuspended solid in 3.0 mL of acetonitrile, 1.0 mL of methanol and 0.2mL of water and stirred at room temperature. As the suspension was tooconcentrated, 2.0 mL of acetonitrile and 4.0 mL methanol were added, andafter about 2 hours an in-process sample was isolated and tested.

Example 18.2 Physical Characterization of the L-Proline Cocrystal ofFormula (1)

¹H NMR spectroscopy of the product from experiment SP221-PRO-P6 revealeda spectrum with signals assignable to proline, which are consistent withthe ratio expected for a cocrystal, although overlapping signalsprevented more detailed quantitative analysis.

The CHNO content of SP221-PRO-P1 was determined by elemental compositionanalysis, with the results shown in Table 45.

TABLE 45 Results from elemental composition analysis of sampleSP221-PRO-P1. Calculated for Calculated for Element % found C₃₆H₄₁N₉O₆C36H41N₉O₆•0.15H₂O C 61.6 62.15 61.91 H 6.3 5.94 5.96 N 18 18.12 18.05 O13.9 13.80 14.09

The PXRD patterns of three samples of this cocrystal, two of which arelikely to be the same crystalline phase (samples SP221-PRO-P1 andSP221-PRO-P3), are shown in FIG. 63. The crystalline phase of samplesSP221-PRO-P1 and SP221-PRO-P3 is designated Form A of L-prolinecocrystal of Formula (1). The following peaks are characteristic of FormA L-proline cocrystal of Formula (1) (as measured for sampleSP221-PRO-P1): 7.0, 10.5, 14.0, 14.8, 17.1, 17.6, 21.1, and 22.1 °2θ±0.2°2θ. PXRD analysis of sample SP221-PRO-P6 (not shown) showed that thismaterial was the same phase as that obtained for sample SP221-PRO-P1.The PXRD pattern of sample SP221-PRO-P2 shows evidence of the presenceof a substantial amount of Form I of the free base of Formula (1).

Raman spectroscopy was measured over the range from 100 cm⁻¹ to 3500cm⁻¹ using a sample of Form A of the L-proline cocrystal of Formula (1)(sample SP221-PRO-P1). The Raman spectrum was obtained in a similarmanner as described in Example 1.2 for Form I. Characteristic Ramanpeaks for Form A of the L-proline cocrystal of Formula (1) are observedat 3064, 2971, 2919, 2881, 2242, 1675, 1607, 1577, 1530, 1484, 1454,1434, 1344, 1323, 1311, 1264, 1243, 1185, 1157, 1035, 1020, 993, 370,275, 190, and 153 (Raman shift, cm⁻¹±2 cm⁻¹).

IR spectroscopy was measured over the range from 600 cm⁻¹ to 3600 cm⁻¹using a sample of Form A of the L-proline cocrystal of Formula (1)(sample SP221-PRO-P1). The IR spectrum was obtained in a similar manneras described in Example 1.2 for Form I. Characteristic IR peaks for FormA of the L-proline cocrystal of Formula (1) are observed at 3471, 3310,3108, 2358, 1672, 1615, 1526, 1495, 1452, 1431, 1399, 1342, 1305, 1262,1241, 1184, 1148, 1091, 1038, 1014, 991, 944, 890, 872, 837, 816, 775,756, 737, 713, and 668 (IR frequency, cm⁻¹±4 cm⁻¹).

The TG-FTIR thermogram of Form A of the L-proline cocrystal of Formula(1) (sample SP221-PRO-P1) was obtained. A small mass loss of about 0.7%observed for sample SP221-PRO-P1 is attributable to water, and no othersolvent loss was observed. The TG-FTIR thermogram of another sample ofForm A of the L-proline cocrystal of Formula (1) (sample SP221-PRO-P3).This sample showed a much larger mass loss, identified as a mixture ofethanol and water by TG-FTIR, which may result from residual solvent.

Example 19 Characterization of the D-Sorbitol Cocrystal of(S)-4-(8-Amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideExample 19.1 Preparation of Form A of the D-Sorbitol Cocrystal ofFormula (1)

A cocrystal of Formula (1) with D-sorbitol was identified in thescreening experiments describe previously. Sample SP221-SBT-P1 wasprepared as follows. 235 mg of Formula (1) free base (PP502-P1) wasdissolved in 12.0 mL of a 0.05 M stock solution of D-sorbitol inmethanol by gentle heating to reflux temperature. The solution wascooled to room temperature and stirred. After several days, no formationof solid was observed, and some of the methanol was evaporated undernitrogen flow. Within three days a suspension was formed, from which thesolid was filtered off and examined by PXRD.

Sample SP221-SBT-P2 was prepared as follows: to the remaining sampleSP22-SBT-P1 was added 3.0 mL of ethanol. The resulting suspension wasstirred at room temperature for four days. The solids were filtered,dried in air at room temperature, and examined by PXRD.

Sample SP221-SBT-P4 was prepared as follows: 510 mg of Formula (1) freebase (PP502-P1) and 200 mg of sorbitol (Sigma #S1876) were dissolved in10.0 mL of ethanol and 3.333 mL of water by gentle heating. The solutionwas cooled to room temperature while purging with a slight nitrogenflow. After three days, a white suspension was obtained from which thesolid was filtered off and dried in air at room temperature for a yieldof about 250 mg. Optical microscopy showed the presence of a very fineneedle shaped crystalline material.

Example 19.2 Physical Characterization of the D-Sorbitol Cocrystal ofFormula (1)

¹H NMR spectroscopy of the product from experiment SP221-SBT-P4 revealeda ratio of Formula (1) free base to D-sorbitol of 3:2.

In addition, sample SP221-SBT-P4 was analyzed for CHNO content byelemental composition analysis, with the results shown in Table 46. Themolecular sum formula of a 1:0.66 cocrystal of Formula (1) andD-sorbitol would be C₂₆H₂₃N₇O₂.0.66[C₆H₁₄O₆] with a molecular weight of586.95 g/mol. A tetrahydrate would be expected to have a molecularweight of 659.0 g/mol (with a water content of 10.9%). The results forC, H, and N are in good agreement with a tetrahydrate, however, theoxygen content found is too low and is more consistent with an anhydrousphase.

TABLE 46 Results from elemental composition (CHNO analysis) and watercontent (TG-FTIR) analysis for sample SP221-SBT-P4. Element % foundC₃₃H₃₁N₇O₇ C 54.8 54.68 H 5.5 6.17 N 15.1 14.88 O 17.2 24.28 water 11.010.9 *This value was taken from the TG-FTIR analysis of SP221-SBT-P4, asdescribed below.

Optical microscopy of the D-sorbitol cocrystal of Formula (1) (sampleSP221-SBT-P4) showed crystalline material with predominantly with veryfine needle-shaped particles with lengths of about 50 to 150 μm andwidths of about 1 to 5 μm.

A PXRD pattern of the D-sorbitol cocrystal of Formula (1) is shown inFIG. 64 (sample SP221-SBT-P4). This crystalline phase is designated FormA of the D-sorbitol cocrystal of Formula (1). The following peaks arecharacteristic of Form A of the D-sorbitol cocrystal of Formula (1):3.0, 4.4, 5.1, 6.0, 7.3, 8.3, 8.6, 10.1, 12.3, 12.7, 13.4, 13.0, 14.3,14.5, 15.6, 16.5, 17.1, 18.0, 18.5, 19.0, 19.4, 19.8, 20.5, 21.2, 21.8,22.3, 22.6, 24.4, 25.5, 27.3, 28.0, and 32.9 °2θ±0.2 °2θ.

Raman spectroscopy was measured over the range from 100 cm⁻¹ to 3500cm⁻¹ using a sample of Form A of the D-sorbitol cocrystal of Formula (1)(sample SP221-SBT-P4). The Raman spectrum was obtained in a similarmanner as described in Example 1.2 for Form I. Characteristic Ramanpeaks for Form A of the D-sorbitol cocrystal of Formula (1) are observedat 3071, 2921, 2246, 1682, 1610, 1579, 1531, 1493, 1453, 1437, 1343,1311, 1246, 1183, 1162, 1039, 1000, 950, and 646 (Raman shift, cm⁻¹±2cm⁻¹).

IR spectroscopy was measured over the range from 600 cm⁻¹ to 3600 cm⁻¹using a sample of Form A of the D-sorbitol cocrystal of Formula (1)(sample SP221-SBT-P4). The IR spectrum was obtained in a similar manneras described in Example 1.2 for Form I. Characteristic IR peaks for FormA of the D-sorbitol cocrystal of Formula (1) are observed at 3207, 2244,1681, 1666, 1651, 1615, 1578, 1548, 1531, 1504, 1463, 1434, 1418, 1398,1311, 1243, 1207, 1149, 1112, 1093, 1052, 1017, 1004, 948, 891, 867,821, 777, 735, 724, 707, 643, and 618 (IR frequency, cm⁻¹±4 cm⁻¹).

The TG-FTIR thermogram of Form A of the D-sorbitol cocrystal of Formula(1) (sample SP221-SBT-P4) indicates a water content of about 11%. Thiswater content is consistent with the expected water content for a 3:2cocrystal of Formula (1):D-sorbitol tetrahydrate for which the watercontent would be 10.9%. The water appears to be weakly bound, becausethe mass loss essentially begins with the beginning of the heatingphase, even from a sample pan with a pinhole as used in the TG-FTIRtesting. Differential scanning calorimetry of Form A of the D-sorbitolcocrystal of Formula (1) (sample SP221-SBT-P4) shows an endotherm at102° C. with a shoulder at 107° C. that may be assigned to the meltingof the cocrystal. The endotherm is followed by exothermic event with apeak temperature of 178° C. that may correspond to recrystallization ofthe free base Form I or thermal degradation.

The results of DVS analysis of Form A of the D-sorbitol cocrystal ofFormula (1) (sample SP221-SBT-P4) show that tested sample can absorb upto about 13% of water at high humidity and that all water is removed at0% RH. A small hysteresis is observed below 20% RH; this suggests theexistence of multiple hydrate forms.

Example 20 Characterization of the Succinic Acid Complex of(S)-4-(8-Amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideExample 20.1 Preparation of Form A of the Succinic Acid Complex ofFormula (1)

Sample SP221-SUC-P4 was prepared as follows: 941 mg of Formula (1) freebase (PP502-P1) and 237.9 mg succinic acid were dissolved in 20.0 mL ofacetone and 3.0 mL of water at 55° C. The solution was cooled to roomtemperature and stirred in an open vial for three days at roomtemperature. After three days the volume of the obtained suspension wasabout 7 mL. The suspension was filtered and the solid dried in air atroom temperature.

Sample SP221-SUC-P1 was prepared as follows: 3.0 mL of stock solution ofFormula (1) (batch SP221-FBL2) was mixed with 3.0 mL stock solution of0.05 M succinic acid in ethanol and allowed to evaporate from an openvial at room temperature. A yellow glassy residue was obtained, to whichwas added 3 mL acetonitrile with continued stirring, after whichevaporation occurred and solids were obtained.

Sample SP221-SUC-P7 was prepared as follows: to 466 mg of Formula (1)free base (batch PP502-P1) and 118 mg of succinic acid were combined in10 mL of 1-propanol. The mixture was heated to 70° C., and 0.1 mL offormic acid was added to achieve dissolution. The solution was allowedto cool to room temperature, and after overnight stirring a suspensionwas obtained from which the solid product was filtered off and dried inair at room temperature. A yield of about 395 mg was obtained.

Sample SP221-SUC-P2 was prepared as follows: 115.5 mg of Formula (1)free base (batch PP502-P1) and 29.0 mg succinic acid were vigorouslyground in an agate mortar with 200 acetone:water 4:1. A sticky paste wasobtained, to which acetonitrile was added; the material remained stickyafter evaporation of the acetonitrile. The mortar was washed out with 15mL of methanol and 2.0 mL of acetonitrile, and the wash solution wasplaced in a glass vial and stirred with the lid open at 40° C. Afterfour days, the suspension was filtered and solids were dried in air atroom temperature.

Preparation of hydrates of cocrystals of Formula (1) and succinic acidmay be achieved by exposing the foregoing organic solvates to water byvapor exchange.

Example 20.2 Physical Characterization of Form A of the Succinic AcidComplex of Formula

¹H NMR spectroscopy of the product from experiment SP221-SUC-P4determined a ratio of Formula (1) free base to succinic acid of about1:1 based on the integral for the aromatic proton of Formula (1) near8.4 ppm and the four protons from the methylene groups of succinic acidbetween 2.4 ppm.

The pKa values of succinic acid are 4.21, and 5.64. Since the lower pKaof succinic acid is just about 1.6 units below the pKa value of basicfunction of ACP-196 free base, co-crystal formation is likely to befavored over salt formation.

A reflection PXRD pattern of a succinic acid cocrystal of Formula (1) isshown in FIG. 65 (sample SP221-SUC-P4). This crystalline phase isdesignated Form A of the succinic acid cocrystal of Formula (1). Thefollowing peaks are characteristic of Form A (acetone solvate) of thesuccinic acid cocrystal of Formula (1): 5.2, 7.5, 8.0, 8.4, 10.3, 12.1,13.3, 14.2, 14.7, 15.5, 16.0, 17.1, 17.5, 18.1, 18.6, 19.2, 19.6, 20.5,21.2, 22.5, 22.8, 23.2, 23.6, 25.5, 26.2, 27.4, 28.2, 28.7, 29.4, 30.3,31.0, 31.6, 32.2, 33.2, and 35.59 °2θ±0.2 °2θ.

Raman spectroscopy was measured over a range from 100 cm⁻¹ to 3500 cm⁻¹using a sample of Form A (acetone solvate) of the succinic acidcocrystal of Formula (1) (sample SP221-SUC-P4). The Raman spectrum wasobtained in a similar manner as described in Example 1.2 for Form I.Characteristic Raman peaks for Form A (acetone solvate) of the succinicacid cocrystal of Formula (1) are observed at 2973, 2922, 2252, 1670,1613, 1580, 1566, 1545, 1529, 1496, 1450, 1347, 1330, 1307, 1270, 1244,1190, 1160, 1036, 1010, 844, 728, 634, 411, 237, 200, and 138 (Ramanshift, cm⁻¹±2 cm⁻¹).

IR spectroscopy was measured over a range from 600 cm⁻¹ to 3600 cm⁻¹using a sample of Form A (acetone solvate) of the succinic acidcocrystal of Formula (1) (sample SP221-SUC-P4). The IR spectrum wasobtained in a similar manner as described in Example 1.2 for Form I.Characteristic IR peaks for Form A (acetone solvate) of the succinicacid cocrystal of Formula (1) are observed at 2359, 1712, 1668, 1620,1603, 1578, 1526, 1494, 1432, 1417, 1403, 1367, 1346, 1304, 1246, 1220,1174, 1162, 1130, 1096, 1035, 1009, 993, 965, 950, 896, 863, 851, 838,778, 754, 734, 726, 712, 672, 636, 624, and 606 (IR frequency, cm⁻¹±4cm⁻¹).

The TG-FTIR thermogram of Form A of the succinic acid cocrystal ofFormula (1) (sample SP221-SUC-P4) shows a significant mass loss ofapproximately 7.5%, identified as acetone, indicates that this sample isan acetone solvate. The TG-FTIR thermogram of the acetonitrile solvateof the succinic acid cocrystal of Formula (1) (sample SP221-SUC-P2),which showed a similar PXRD pattern compared to sample SP221-SUC-P1. Amass loss of nearly 4%, attributable mainly to acetonitrile, indicatesthat this sample is an acetonitrile solvate. The TG-FTIR thermogram ofthe acetonitrile solvate of the succinic acid cocrystal of Formula (1)(sample SP221-SUC-P7) shows a mass loss of nearly 9.5%, attributablemainly to 1-propanol, indicates that this sample is a 1-propanolsolvate.

Example 21 Characterization of the Sulfate Salt of(S)-4-(8-Amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamideExample 21.1 Preparation of Form A of the Sulfate Salt of Formula (1)

A sulfate salt of Formula (1) (sample SP221-SO4-P1) was prepared asfollows. To 5.0 mL of a 0.1 M stock solution of Formula (1) free base inacetone-water (sample SL20150415FB, 0.1 M) was added one equivalent ofsulfuric acid in form of concentrated sulfuric acid (27.8 μL), which washeated to 50° C. and allowed to cool to room temperature. Ascrystallization did not occur the mixture was seeded with a few mg ofcrystalline phosphate salt. After overnight stirring at room temperaturea yellow/white suspension was obtained from which the solid was filteredoff and dried in air at room temperature.

Sample SP221-SO4-P3 was prepared by repetition of the experiment used toproduce SP221-SO4-P1 using a 1:1 ratio of sulfuric acid to free base.

Sample SP221-SO4-P4 was prepared by dissolving 941 mg of sample PP502-P1in 22 mL of acetone-water 10:1 at about 50° C. and adding one equivalentof concentrated sulfuric acid (112 μL). A suspension formed at 50° C.;the mixture was allowed to cool to room temperature and then stirredovernight at room temperature before solid was filtered off and dried inair. About 880 mg of slightly yellowish solid was obtained.

Sample SP221-SO4-P5 was prepared by adding 300 mg of SP221-SO4-P4 to 3.0mL of acetonitrile and 0.3 mL of water. The suspension was stirred atroom temperature for one day. The suspension was filtered, and solidswere dried in air at room temperature.

Sample SP221-SO4-P6 was prepared by adding 944 mg of Formula (1) freebase (batch PP502-P1) to 15.0 mL of acetone:water (9:1) and heating toreflux to achieve dissolution. Sulfuric acid (0.8 mL/1 equivalent) wasthen added in the form of a 2.5 M aqueous solution. The solution wasseeded with SP221-SO4-P1 and allowed to cool to 35° C. while stirringwas continued overnight. A suspension was obtained that was reheated to50° C. for about three hours then allowed to cool again to roomtemperature and stirred for two hours before the solid was filtered offand dried in air at room temperature. A yield of about 950 mg wasobtained.

Example 21.2 Physical Characterization of Form A of the Sulfate Salt ofFormula (1)

¹H NMR spectroscopy (spectrum not shown) of the product from experimentSP221-504-P4 was consistent with Formula (1). Samples SP221-SO4-P4 andSP221-SO4-P5 were analyzed by CHONS elemental composition analysis, withthe results shown in Table 47. The molecular sum formula expected for asolvent-free monostoichiometric sulfate salt of Formula (1) isC₂₆H₂₅N₇O₆S, with a molecular weight of 563.6 g/mol. A trihydratemonostoichiometric sulfate salt of Formula (1) is expected to have a summolecular formula of C₂₆H₃₁N₇O₉S and a molecular weight of 617.6 g/mol(with a water content of 8.7%). A tetrahydrate monostoichiometricsulfate salt of Formula (1) is expected to have a molecular sum formulaof C₂₆H₃₃N₇O₁₀S and a molecular weight of 635.7 g/mol (with a watercontent of 11.3%). The best fit to the experimental values for sampleSP221-SO4-P4 is to a tetrahydrate with an excess of sulfuric acidequivalent to a molar ratio of about 1.25. The best fit to theexperimental values for sample SP221-SO4-P5 was found for a monosulfatetetrahydrate salt, wherein good agreement with the theoreticallyexpected hydrogen, oxygen and sulfur content was found, with only aslight discrepancy found for carbon and nitrogen.

TABLE 47 Results from elemental composition (CHNOS analysis) analysisfor samples SP221-SO4-P4 and SP221-SO4-P5 of the sulfate salt of Formula(1). SP221-SO4-P4 (% SP221-SO4-P5 (% C₂₆H_(33.5)N₇O₁₁S_(1.25)Tetrahydrate found, found, tetrahydrate sulfate monosulfate salt Elementexperimental) experimental) salt (theoretical) (theoretical) C 47.4 46.847.30 49.13 H 4.9 4.7 5.11 5.23 N 14.9 14.4 14.85 15.42 O not determined25.0 26.66 25.17 S 6.2 5.0 6.07 5.04 water* 10.6 not available 10.9 11.3*Water content was determined by Karl Fischer titration.

Optical microscopy of sample SP221-SO4-P5 showed crystalline materialwith predominantly needle-shaped particles. The particles in sampleSP221-SO4-P5 were considerably smaller than those for sampleSP221-SO4-P6, which showed particle lengths up to about 100 μm andwidths of about 5 to 10 μm.

The reflection PXRD pattern of sample SP221-SO4-P6 is shown in FIG. 66.This crystalline phase is designated Form A of the sulfate salt ofFormula (1). The following peaks are characteristic of Form A of thesulfate salt of Formula (1): 4.6, 5.0, 8.0, 9.0, 9.8, 12.0, 12.7, 13.2,14.6, 15.0, 15.6, 16.2, 17.5, 18.0, 19.8, 20.2, 21.9, 23.8, 24.4, 24.9,25.7, 26.0, 27.2, 29.5, 30.4, 31.6, and 32.5 °2θ±0.2 °2θ.

Raman spectroscopy was measured over a range from 100 cm⁻¹ to 3500 cm⁻¹using a sample of Form A of the sulfate salt cocrystal of Formula (1)(sample SP221-SO4-P4). The Raman spectrum was obtained in a similarmanner as described in Example 1.2 for Form I. Characteristic Ramanpeaks for Form A of the sulfate salt of Formula (1) are observed at3115, 2977, 2926, 2224, 1675, 1611, 1537, 1498, 1449, 1409, 1361, 1327,1310, 1288, 1243, 1198, 1155, 1042, 1009, 978, 948, 906, 849, 771, 713,652, 632, 464, 370, and 254 (Raman shift, cm⁻¹±2 cm⁻¹).

IR spectroscopy was measured over a range from 600 cm⁻¹ to 3600 cm⁻¹using a sample of Form A of the sulfate salt of Formula (1) (sampleSP221-SO4-P4). The IR spectrum was obtained in a similar manner asdescribed in Example 1.2 for Form I. Characteristic IR peaks for Form Aof the sulfate salt of Formula (1) are observed at 3430, 3101, 3029,2225, 1667, 1633, 1615, 1598, 1563, 1557, 1508, 1428, 1350, 1328, 1308,1276, 1225, 1088, 1036, 1018, 925, 891, 848, 816, 783, 736, 723, 694,and 612 (IR frequency, cm⁻¹±4 cm⁻¹).

The TG-FTIR thermogram of Form A of the sulfate salt of Formula (1)(sample SP221-504-P4) shows a mass loss of 10.1% is due to water. Thewater loss begins with heating and is complete by about 110° C. using aheating rate of 10° C. per minute. Differential scanning calorimetryresults for Form A of the sulfate salt of Formula (1) (sampleSP221-SO4-P4). DSC showed a melting endotherm with a peak temperature of118° C. and an enthalpy of fusion of about 92 J/g. DSC revealed amelting endotherm at 127° C. with an enthalpy of fusion of about 70 J/g.

DVS analysis was performed for Form A of the sulfate salt of Formula (1)(sample SP221-SO4-P4) show that the water is not completely removed at0% RH after five hours. The initial water content of the given sulfatesample was determined by Karl Fischer titration. At the end of the test,the water content is almost the same as at the beginning of themeasurement.

Example 22 Overcoming the Effects of Acid Reducing Agents withFormulations of(S)-4-(8-Amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide

Acid-reducing agents, such as omeprazole, may limit the exposure ofFormula (1) free base in mammals because of previously-discussedpH-solubility profile of Formula (1). This is a significant issue in thetreatment of patients with cancer, inflammatory diseases, immunediseases, and autoimmune diseases, since these patients are commonlyco-administered acid reducing agents for gastric irritation that oftenaccompanies their conditions. Acid reducing agents are the most commonlyprescribed medications in North America and Western Europe. Of recentlyapproved oral cancer therapeutics, >50% have pH-dependent solubility,and therefore have a potential drug-drug interaction with regards toacid reducing agents. In cancer patients, it is estimated that 20-33% ofall patients are using some form of acid-reducing agent. In particularcancers, such as pancreatic cancer or gastrointestinal cancers, acidreducing agent use is as high as 60-80% of patients. Smelick, et al.,Mol. Pharmaceutics 2013, 10, 4055-4062.

The concern of potential drug-drug interactions with acid reducingagents for weakly basic drugs has led to the development of riskassessment strategies and drug-drug interaction studies designs for newdrugs that exhibit pH dependent solubility and dissolution. Smelick, etal., Mol. Pharmaceutics 2013, 10, 4055-4062. Acid reducing agentsinclude proton pump inhibitors, such as omeprazole, esomeprazole,lansoprazole, dexlansoprazole, pantoprazole, rabeprazole, andilaprazole; H₂ receptor antagonists, such as cimetidine, ranitidine, andfamotidine; and antacids such as bicarbonates, carbonates, andhydroxides of aluminium, calcium, magnesium, potassium, and sodium.Mixtures of antacids plus agents targeting mechanisms of gastricsecretion may also be used as prescription or non-prescriptionacid-reducing agents. Any other acid reducing agent known in the art mayalso be used. In some cases, the effect of an acid-reducing agent istransient and depends on presence of the agent in the stomach. In othercases, the effect of an acid-reducing agent may be pronounced throughoutthe treatment interval, providing a constant elevation of gastric pH tolevels greater than pH 4.

The terms hypochlorhydria and achlorhydria refer to conditions wheregastric secretion of hydrochloric acid is lower than normal or severelyreduced to nonexistent. The natural pH of the stomach is lowered by acidsecretions in response to food stimulation; in certain medicalconditions the ability of the gastric proton pump to secrete acid iscompromised. Infections with H. pylori have been associated withimpaired secretion of gastric acid (hypochlorhydria or achlorhydria).Other disease states, including those in which gastric parietal cellsare destroyed or depleted, or the signaling to gastric parietal cells isaltered, can lead to hypochlorhydria or achlorhydria. Long-term use ofproton pump inhibitors or H₂ receptor antagonists may also result inthese conditions. The evaluation of gastric pH over the course of a day(through meals) may be monitored in patients with in situ pH probes, ifneeded, as a diagnostic aid.

Dissolution of Form 1 of Formula (1) into aqueous media such as stomachfluid is pH dependent (see, e.g., FIG. 67 and FIG. 68, discussed in moredetail below). The bioavailability of Formula (1) may therefore bemodified by factors that improve its dissolution. Alternate forms ofFormula (1), and acidification of the formulation of Form 1 of Formula(1) were tested in dogs treated with omeprazole 10 mg/day to evaluatethe extent to which an alternate form of Formula (1) can overcome theeffects of acid-reducing agents.

Dogs were treated with Formula (1) 100 mg capsules in several relatedstudies using the same animals and a strict dosing schedule to minimizeintra- and inter-animal variability. All doses were chased with 35 mL ofdistilled H₂O by oral gavage to standardize the dissolution volume witheach dose administration. Dogs were conditioned to receive sham capsulesand chase water on non-dosing days; food was also controlled to reducethe variability associated with gastric acid secretions in response topresentation and consumption of chow. The conditioning regimen wasfollowed continuously for at least six months, and the same 12 dogs wereused for all studies described below.

Study 2219-057 used 100 mg of Formula (1) in liquid capsules(hydroxyl-β-cyclodextrin/citrate, 2 doses) to set the bar for absorptionwithout a dissolution component associated with solid form. Study2219-059 used Formula (1) with formulation F-1 and Study 2219-061 usedFormula (1) with formulation F-2 alone or following pre-treatment of thedogs with omeprazole, then proceeded to test Formula (1) salt forms inthe F-1 formulation and an acidic formulation of Form I of Formula (1)designated FA-3 (see Example 23 below for the preparation offormulations). FIG. 69 shows individual concentration-time profiles forall dogs included in this series of studies. For simplicity, the meanconcentration-time profiles are presented in FIG. 69 for studies withrepeated doses of the same form/formulation of Formula (1). The Y axesin FIG. 69 are set for each dog individually, to emphasize thewithin-dog effects of salt forms and prototype acid formulation.

Study 2219-061 used Form I of Formula (1) recrystallized from ethanol,as described in Example 8, and the maleate, phosphate, fumarate andtartrate salt forms of Formula (1). Following collection ofpharmacokinetics data after one dose administration of 100 mg capsulesin the F-2 formulation, omeprazole treatment (10 mg/day) was initiatedas a part of the conditioning regimen. The remaining study phases wereconducted in omeprazole treated dogs. After 4 days on omeprazole, thedogs were dosed with experimental Formula (1) drug forms or formulationon top of the continuing daily omeprazole dose. Salt forms were dosed toequal 100 mg of Formula (1) free base. The F-1 formulation was used fordosing the salt forms after correction for counterion and water content.The prototype acid formulation (FA-3) used both fumaric acid and alginicacid as an extra-granular mix with the granulated Formula (1) informulation F-2.

FIG. 70 shows changes in the AUC, C_(max) and T_(max) by study phase,with each study or study phase presented sequentially. The initial studyusing liquid formulation in capsules to deliver 100 mg Formula (1) insolution was designed to show exposures following this dose (i.e., notdissolution limited), and to characterize the variability in dogs whendissolution-associated variance is removed. The higher mean exposure andsmaller within- and between-animal variability observed followingadministration of the fully dissolved Formula (1) in liquid capsules toconditioned dogs demonstrate that dissolution of Form I of Formula (1)plays a role in limiting oral absorption, indicating that optimaldissolution will enhance absorption.

The remaining variance in pharmacokinetic parameters observed followingadministration of liquid capsules may be due to intrinsic factors thatvary between the inbred beagle dogs. This effect has also beendemonstrated using 25 mg liquid capsules and Form I of Formula (1)capsules using a dose scaled version of F-1. Between-animal variabilityfollowing administration of a liquid capsule or matching solid capsulescontaining Form 1 of Formula (1) at a fixed dose, may result from smallvariations in the mg/kg dosage of test article as well as otherintrinsic factors such as those governing drug metabolism andelimination. Adding a weight-based normalization for the AUC's in thesolid form experiments with this group of dogs will further tighten thebetween-animal variance at each dosing interval. Dose adjusted AUC andC_(max) values can be compared most accurately for statistical analysisof the experimental results.

After administration of the salt forms or Form I of Formula (1) inacidic formulation to overcome the omeprazole effect, T_(max) increasedin most of the dogs. Although there was a trend towards a lower meanC_(max) in these study phases (FIG. 69 and FIG. 70), the pattern was notobserved with every phase or for all dogs. A similar trend has beenobserved in dogs and humans when Form I of Formula (1) is administeredwith food. Notably, the mean AUC levels in dogs treated with salt formsof Formula (1) or with the acidic formulation of Form I, were similar toAUCs observed in dogs after administration of Form I without omeprazole.There was a trend to decrease between-animal variability when theseexperimental dosage forms were administered, compared with the Form Icapsules administered in conditioned dogs without omeprazole treatment.Therefore, exposure after oral dosing with Formula (1) in a salt formare increased in the presence of omeprazole, and variability in exposureis decreased in both omeprazole-treated dogs and conditioned dogswithout omeprazole treatment. The prototype acidic formulation for FormI of Formula (1) has a similar effect.

The observed effect of alternate salt forms and acidulents with Formula(1) on oral absorption in omeprazole-treated dogs is novel andsurprising. The pH-dependence of Formula (1) dissolution is associatedwith the stability of acidic and basic species in aqueous solutions, andwith the free energy of dissolution during phase transition. In vitro-invivo correlations demonstrated that dissolution limitations wereassociated with poor absorption of Form I of Formula (1) inomeprazole-treated dogs (or dogs treated with alternate gastric acidreducing agents, such as famotidine, calcium carbonate, or the othertreatments listed above). In a human Phase 1, single-center, open-label,fixed-sequence, 2-period, 3-part study to evaluate the one-wayinteraction of calcium carbonate, omeprazole, or rifampin on Formula (1)in healthy adult subjects, treatment of subjects with acid reducingagents prior to administration of Form I of Formula (1) resulted insignificant decreases in exposure. The role of pH in dissolution ofFormula (1) was demonstrated in vitro, and the dissolution limitation onabsorption was postulated in vivo. The addition of acidulants to theformulation, or the administration of fully dissolved Formula (1), aremethods to facilitate dissolution by lowering pH in themicroenvironment, or to circumvent the dissolution step for aproof-of-concept in vivo model. In contrast, dosing with the alternatesalt forms of Formula (1), which were expected to have little impact ongastric pH, demonstrated that the solid form of Formula (1) hassignificant and unexpected effects on oral absorption characteristics inmammals.

FIG. 71 compares dose-normalized AUC and C_(max), with additionalaveraging of repeated exposures for dogs for liquid capsules (n=2 perdog) and F-2 (n=5 per dog). The results again show that Formula (1)exposure can be recovered in the presence of omeprazole using the FA-3acidulant formulation of the present invention as well as the salts ofthe present invention.

These studies demonstrate that good exposures can be achieved inomeprazole treated dogs via either an enabling formulation for Form I ofFormula (1), or by generating a new salt form of Formula (1). Theexposures obtained using formulation FA-3 with acidulant and the saltswith omeprazole are surprisingly similar to exposures observed withoutomeprazole, and would be expected to perform well for other salts andacidulants as well as for other acid-reducing agents. Thedissolution-mediated absorption observed in human subjects can bemodeled in dogs. The in vitro dissolution assay is also a good predictorof in vivo absorption characteristics of different encapsulatedformulations of Form I of Formula (1).

A separate PK comparability study (2219-060) has been completed tocharacterize exposures from the acetone-recrystallized andethanol-recrystallized drug substance in capsules manufactured with theF-2 formulation. These data more fully characterize the between- andwithin-dog variability associated with absorption of Formula (1) inconditioned dogs that are not treated with omeprazole and indicate thatethanol-recrystallized drug substance is suitable for late-phaseclinical development.

Example 23 Formulations of(S)-4-(8-Amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide

Formulations of Formula (1) solid forms (salts and free base Form I)were prepared as shown in Table 48.

TABLE 48 Description of Formulation 1 (F-1) and acidic saltformulations. F-1 F-1 F-1 F-1 F-1 free base maleate fumarate L-tartratephosphate (Form I) (Form A) (Form A) (Form A) (Form A) w/w % w/w % w/w %w/w % w/w % Formula (1) 10-50% 10-50% 10-50% 10-50% 10-50% Micro- 50-90%50-90% 50-90% 50-90% 50-90% crystalline Cellulose

Additional formulations were prepared as shown in Table 49 using thefollowing procedure. Formula (1) dry-granulate was blended withextragranular acidulants. The blends were then filled into hard gelatincapsules (in the case of FA-1, FA-2, FA-4, and FA-5) or compressed intoa tablet (in the case of FA-4).

TABLE 49 Formulations of Formula (1), showing intragranular componentsand extragranular components. SILICIFIED PARTIALLY SODIUM FORMU-MICROCRYSTALLINE PRE-GELATINIZED STARCH MAGNESIUM LA (1) CELLULOSE MAIZESTARCH GLYCOLATE STEARATE F-2 w/w % Intra- 25-50% 25-33% 20-33% 0-5%0.05-1% FA-1 w/w % granular 25-35% 15-35% 20-33% 0-5% 0.05-1% FA-2 w/w %25-35% 15-35% 20-33% 0-5% 0.05-1% FA-3 w/w % 25-35% 15-35% 20-33% 0-5%0.05-1% FA-4 w/w % 25-35% 15-35% 20-33% 0-5% 0.05-1% FA-5 w/w % 25-35%15-35% 20-33% 0-5% 0.05-1% MAGNESIUM FUMARIC ALGINIC POLOXAMERHYDROXYPROPYL STEARATE ACID ACID 407 METHYLCELLULOSE F-2 Extra- 0.05-1%— — — — FA-1 granular 0.05-1% 25-33% — — 1-5% FA-2 0.05-1% — 15-33%  — —FA-3 0.05-1% 15-33% 5-15% — — FA-4 0.05-1% 15-33% 5-15% — — FA-5 0.05-1%15-33% 5-15% 0.5-5% — 1. Polaxamer 407 refers to triblock copolymer ofpolypropylene glycol and polyethylene glycol (available from BASF, Inc.,under the tradename PLURONIC F127).

The results of dissolution experiments using formulations representativeof those in Table 49 at two different pH values are shown in FIG. 67 andFIG. 68. The dissolution system was a U.S. Pharmacopeia Type IIapparatus equipped with paddles (at 50 rpm) and 900 mL vesselsequilibrated at 37° C. Samples are taken at intervals using a cannula ata set depth through an in-line filter and analyzed by reversed phaseHPLC with UV spectroscopic detection. Capsules were tested in sinkers,and the tablet was tested neat.

Additional formulations were prepared according to Table 50.Intragranular formulations may be prepared by the following procedure.Materials are pre-blended in a 250 mL V-blender for 300 revolutions.After blending, lubricant is added and blending is performed for 100additional revolutions. The blend is roller compacted on a TF-miniroller compactor and then feed through an oscillating granulatorequipped with a 20 mesh screen. Extragranular formulations may beprepared by the following procedure. When extragranular acids orpolymers are added, they are added to the preblended or neatextragranular material and then add granulated in a 250 mL V-blender andfor 300 revolutions. After blending, lubricant is added and blending isperformed for an additional 100 revolutions. Lubricated granules arethen filled into size 1 hard gelatin capsules using a dosing disk ordosator-equipped semi-automatic or automatic encapsulator. Alternately,material may be compressed on a tablet press or mold.

TABLE 50 Formulations of Formula (1). SILICIFIED PARTIALLY SODIUM FORMU-MICROCRYSTALLINE PRE-GELATINIZED STARCH TARTARIC MAGNESIUM LA (1)CELLULOSE MAIZE STARCH GLYCOLATE ACID STEARATE FA-7 w/w % Intra- 25-50%25-33% 20-33% 0-5% 15-33% 0.05-1% FA-8 w/w % granular 25-35% 15-35%20-33% 0-5% — 0.05-1% FA-9 w/w % 25-35% 15-35% 20-33% 0-5% — 0.05-1%FA-10 w/w % 25-35% 15-35% 20-33% 0-5% — 0.05-1% ALGINIC MAGNESIUMTARTARIC ALGINIC ASCORBIC CARBOPOL HYDROXYPROPYL ACID STEARATE ACID ACIDACID 971P¹ METHYLCELLULOSE FA-7 5-15% Extra- 0.05-1% — — — — — FA-8 —granular 0.05-1% 25-33%  5-15% — — — FA-9 — 0.05-1% — 15-33% — — 7.5-15%FA-10 — 0.05-1% — — 20-50% 2.5-10% — ¹Carbopol 971P (Lubrizol, Inc.) isalso referred to as carboxypolymethylene or “carbomers” (e.g., inpharmaceutical monographs such as USP/NF or Ph. Eur).

In addition to the formulations described in Table 49 and Table 50,other acidulants may also be used as described herein, including fumaricacid, succinic acid, D-tartaric acid, L-tartaric acid, racemic tartaricacid, ascorbic acid, isoascorbic acid (also known as erythorbic acid andD-araboascorbic acid), alginic acid, Protacid F 120 NM, Protacid AR 1112(also known as Kelacid NF), Carbopol 971P (carboxypolymethylene), andCarbomer 941 (polyacrylic acid).

Additional non-limiting formulations are given in Table 51, and may beprepared as described above or by using methods known in the art. Theseformulations, and all of the foregoing formulations, may be prepared ascapsules or tablets, with or without coating.

TABLE 51 Formulations of Formula (1). FORMULA (I) SILICIFIED PARTIALLYSODIUM (FORM I FREE MICROCRYSTALLINE PRE-GELATINIZED STARCH TARTARICBASE) CELLULOSE MAIZE STARCH GLYCOLATE ACID FA-11 w/w % Intra- 25-50%10-25% 10-25% 0-5% 10-30% FA-12 w/w % granular 25-50% 10-25% 10-25% 0-5%— SODIUM MAGNESIUM ALGINIC HYDROXYPROPYL MAGNESIUM TARTARIC STARCHSTEARATE ACID METHYLCELLULOSE STEARATE ACID GLYCOLATE FA-11 0.05-1%10-30% — Extra- 0.05-2% — 0-6% FA-12 0.05-1% — 0-20% granular 0.05-2%10-35% 0-6%

Example 24 Amorphous Dispersions of(S)-4-(8-Amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide

The feasibility of the formation of amorphous solid dispersions ofFormula (1) in following polymers was explored in a variety of polymers,including KOLLIDON VA64, polyvinylpyrollidone (PVP-10 and PVP-40), HPMC,Kolliphor P188, HPMCAS-L, HPMCAS-H, EUDRAGIT L100-5, and EUDRAGIT L100.Drug substance concentrations in the amorphous solid dispersions weregenerally about 11 to 20%. Aqueous solubilities of the amorphous soliddispersions were measured at pH ˜6.8 in phosphate buffer. Typicalsolubilities were between 100 μg/mL and 160 μg/mL, which is about twicethe solubility of Form I of the free base of Formula (1), but less thanthe solubility of the amorphous form of the free base of Formula (1).The polymers used for the preparation of amorphous solid dispersions aresummarized in Table 52.

TABLE 52 Polymers used for amorphous solid dispersions. Polymer SupplierLot Number Vinylpyrrolidone-vinyl acetate copolymer (PVP-VA) BASF50347977 (KOLLIDON VA64) Polyvinylpyrrolidone (PVP-10, average molecularSigma Aldrich BCBF4168V weight 10 kDa) Polyvinylpyrrolidone (PVP-40,average molecular Sigma Aldrich BCBG0598V weight 40 kDa)Hydroxypropylmethylcellulose (HPMC) Sigma Aldrich BCBC9084V Kolliphor(Poloxamere) P188 Sigma Aldrich SLBJ6719V Hydroxypropylmethylcelluloseacetate succinate, Shin-Etsu 3093208 HPMCAS-LF (AQOAT AS-LF)¹Hydroxypropylmethylcellulose acetate succinate, Shin-Etsu 4073131HPMCAS-HF (AQOAT AS-HF)² Methacrylic acid:ethyl acrylate copolymer (1:1)Evonik B141104042 (EUDRAGIT L100-55) Methacrylic acid:methylmethacrylate copolymer (1:1) Evonik B14003011 (EUDRAGIT L100) ¹LF graderefers to HPMCAS polymer with an acetyl content of 5 to 9%, a succinoylcontent of 14 to 18%, a methoxyl content of 20 to 24%, and ahydroxypropyl content of 5 to 9%. ²HF grade refers to HPMCAS polymerwith an acetyl content of 10 to 14%, a succinoyl content of 4 to 8%, amethoxyl content of 22 to 26%, and a hydroxypropyl content of 6 to 10%.

Batch PP502-P111 was prepared as follows: 1.008 g of PVP-K25 (BASF) wasdissolved in water. To 2.0 mL of the PVP solution was added 24 mg ofFormula (1) free base (batch PP502-P1), and dissolution was achieved byadditionally adding 2.0 mL of water, 0.2 mL of acetone, and 0.2 mL ofacetic acid. A clear solution obtained, from which the solvents wereallowed to evaporate at room temperature.

Batch PP502-P112 was prepared as follows: 6.0 mL of THF was added to theremaining PVP-K25 (form BASF) solution from experiment PP502-P111 (18mL). An aliquot of 3 mL of this new solution was taken, and 33 mg ofFormula (1) free base batch PP502-P1 was added to this aliquot. Fulldissolution was not achieved, and 0.1 mL of acetic acid was added. Aclear solution was then obtained, from which the solvents were allowedto evaporate at room temperature.

Batch PP502-P116 was prepared as follows: 2.007 g of PVP-10(Sigma-Aldrich/CR-1104921) was dissolved in 20 mL of water and 202.9 mgof Formula (1) free base batch PP502-P1 was separately dissolved in 4.3mL of acetone:water (4:1). The two solutions were combined and a newclear solution was obtained that was subjected to freeze drying. Aslightly yellow powder was obtained.

Batch PP502-P119, a 10% w/w dispersion of Formula (1) in Kolliphor P188,was prepared as follows: 1.999 g Kolliphor P188 (poloxamer P188,obtained from Sigma-Aldrich #K4894) was dissolved in 20 mL of water, and240 mg of PP502-P1 was separately dissolved into 9.5 mL of acetone:water(4:1). The solution containing Formula (1) was filtered into the polymersolution and the mixture was freeze dried. A solid product was obtainedafter freeze drying.

Batch PP502-P128, a 16% w/w dispersion of Formula (1) in PVP-VA(KOLLIDON VA64), was prepared as follows. Stock solution SL20150715 wasfirst prepared by dissolving 1.532 g of batch PP502-P1 of Formula (1)free base in 33.0 mL of acetone:water (4:1), yielding a solution with aconcentration of 45 mg/mL of Formula (1). 1.01 g of PVP-VA was dissolvedin 10 mL of water, to which was added 4.3 mL of stock solutionSL20150715 (292 mg, to achieve 11% w/w drug loading). A solution wasobtained and freeze dried to yield solid product.

Batch PP502-P129, a 16% w/w dispersion of Formula (1) in PVP10, wasprepared as follows: 999 mg of PVP-10 was dissolved in 10 mL of water,to which was added 4.3 mL of the aforementioned stock solutionSL20150715 (193 mg, to achieve 16% w/w drug loading). A solution wasobtained and freeze dried to yield solid product.

Batch PP502-P130, a 16% w/w dispersion of Formula (1) in PVP40, wasprepared as follows: 1.01 g of PVP-40 was dissolved in 10 mL of water,to which was added 4.3 mL of the aforementioned stock solutionSL20150715 (193 mg, to achieve 16% w/w drug loading). A solution wasobtained and freeze dried to yield solid product.

Batch PP502-P131, an 11% w/w dispersion of Formula (1) in PVP-VA andHPMC, was prepared as follows: 1.02 g of PVP-VA and 506 mg HPMC (for a2:1 polymer ratio) were dissolved in 10 mL of water, to which was added6.5 mL of SL20150715 (292 mg, to achieve 11% drug loading). A solutionwas obtained and freeze dried to yield solid product.

Batch PP502-P136, a 17% w/w dispersion of Formula (1) in EUDRAGITL100-55, was prepared as follows. 1009 mg of Eudragit L100-55 and 201 mgof PP502-P1 (16.6%) were placed in a 250 mL round-bottom flask and weredissolved in 14.0 mL of THF:water (4:1). A yellow-colored solution wasobtained. The THF was evaporated in a rotary evaporator at 38° C., andthen the remaining solution was freeze dried until the next day at apressure of 0.09 mbar. Part of the obtained solid material was isolatedas batch PP502-P136. The remaining part of this material was furtherdried at 60° C. under vacuum for about 18 hours, and was designatedbatch PP502-P136A.

Batch PP502-P137, a 17% w/w dispersion of Formula (1) in EUDRAGIT L100,was prepared as follows: 1011 mg of Eudragit L100 and 200 mg of PP502-P1(16.6%) were placed in a 250 mL round-bottom flask and were dissolved in20.0 mL of THF:water (4:1). A yellow-colored solution was obtained. TheTHF was evaporated in a rotary evaporator at 38° C. and then theremaining solution was freeze dried until the next day at a pressure of0.09 mbar. Part of the obtained solid material was isolated as batchPP502-P137. The remaining part of this material was further dried at 60°C. under vacuum for about 18 hours, and was designated batchPP502-P137A.

Batch PP502-P138, a 20% w/w dispersion of Formula (1) in HPMCAS-LF, wasprepared as follows: 998 mg of HPMCAS-LF and 250 mg of PP502-P1 (20%)were placed in a 250 mL round flask and dissolved in 25.0 mL ofTHF:water (4:1). A yellow-colored solution was obtained. The THF wasevaporated in a rotary evaporator at 38° C., and then the remainingsolution was freeze dried until the next day at a pressure of 0.09 mbar.Part of the obtained solid material was isolated as batch PP502-P138.The remaining part of this material was further dried at 60° C. undervacuum for about 18 hours, and was designated batch PP502-P138A.

Batch PP502-P139, a 20% w/w disperion of Formula (1) in HPMCAS-HF, wasprepared as follows: 1036 mg of HPMCAS-HF and 250 mg of PP502-P1 (20%)were placed in a 250 mL round flask and dissolved in 25.0 mL ofTHF:water (4:1). A yellow-colored solution was obtained. The THF wasevaporated in a rotary evaporator at 38° C., and then the remainingsolution was freeze dried until the next day at a pressure of 0.09 mbar.Part of the obtained solid material was isolated as batch PP502-P139.The remaining part of this material was further dried at 60° C. undervacuum for about 18 hours, and was designated batch PP502-P139A.

PXRD results for several batches are given in FIG. 72. The lack of anydiscernable Bragg reflections in each PXRD pattern demonstrates a lackof detectable crystalline content in each sample, and indicates thateach sample is amorphous. DSC results are summarized in Table 53. Theappearance of a single T_(g), rather than separate T_(g) events for thepolymer and the drug, indications formation of an amorphous solidmolecular dispersion. TG-FTIR results are also summarized in Table 53.

TABLE 53 Summary of DSC and TG-FTIR results for amorphous soliddispersions. Sample identifier Dispersion DSC result (T_(g) and ΔC_(p))TG-FTIR result PP502-P128 16% Formula (1) in T_(g) ~115° C., ΔC_(p)~0.72 J/g/K¹ Δm = −3.8% at 160° C. attributable to PVP-VA T_(g) ~111°C., ΔC_(p) ~0.48 J/g/K² water, no organic solvent PP502-P129 16% Formula(1) in T_(g) ~126° C., ΔC_(p) ~0.99 J/g/K¹ Δm = −4.2% at 140° C.,attributable to PVP-10 T_(g) ~110° C., ΔC_(p) ~0.48 J/g/K² water, noorganic solvent, at higher temperature degradation possible PP502-P13016% Formula (1) in T_(g) ~153° C., ΔC_(p) ~0.57 J/g/K¹ Δm = −5.2% at140° C., attributable to PVP-40 T_(g) ~152° C., ΔC_(p) ~0.37 J/g/K²water, no organic solvent, at higher temperature degradation possiblePP502-P131 11% Formula (1) in T_(g) ~120° C., ΔC_(p) ~0.63 J/g/K¹ Δm =−3.1% at 140° C., attributable to PVP-VA:HPMC(2:1) T_(g) ~114° C.,ΔC_(p) ~0.53 J/g/K² water, no organic solvent, at higher temperaturedegradation possible PP502-P136A 17% Formula (1) in T_(g) ~134° C.,ΔC_(p) ~0.55 J/g/K Δm = −~3.5% at 160° C., attributable EUDRAGIT L100-55to THF PP502-P137A 17% Formula (1) in T_(g) ~143° C., ΔC_(p) ~0.39 J/g/KΔm = −~3.5% at 160° C., attributable EUDRAGIT L100 to THF PP502-P138A20% Formula (1) in T_(g) ~114° C., ΔC_(p) ~0.48 J/g/K Δm = −~2.5% at160° C., THF and HPMCAS-LF water PP502-P139A 20% Formula (1) in T_(g)~111° C., ΔC_(p) ~0.49 J/g/K Δm = −~1.5% at 160° C., attributableHPMCAS-HF to water ¹Measured in closed sample pan. ²Measured in opensample pan.

Additional amorphous solid dispersions with different concentrations ofFormula (1) can also be produced using the procedures described above.

The apparent aqueous solubility of selected amorphous solid dispersionswas tested in phosphate buffer at pH 6.8, with the results shown inTable 54.

TABLE 54 Summary of solubility testing results for amorphous soliddispersions. Solu- Solubil- bility pH ity (24 pH Sample (1 hour) (1hours) (24 identifier Dispersion (mg/ml) hour) (mg/ml) hours) PP502-P12816% Formula (1) in 0.157 6.79 0.105 6.78 PVP-VA PP502-P129 16% Formula(1) in 0.110 6.80 0.07 6.78 PVP-10 PP502-P130 16% Formula (1) in 0.1306.80 0.09 6.80 PVP-40 PP502-P131 11% Formula (1) in 0.160 6.82 0.0886.81 PVP-VA:HPMC (2:1) PP502-P136 17% Formula (1) in 0.018 6.49 0.0156.39 EUDRAGIT L100-55 PP502-P137 17% Formula (1) in 0.059 6.71 0.0436.59 EUDRAGIT L100 PP502-P138 20% Formula (1) in 0.106 6.49 0.091 6.34HPMCAS-LF PP502-P139 20% Formula (1) in 0.100 6.54 0.135 6.58 HPMCAS-HF

Example 25 Comparison of Processability for Free Base Form I and FreeBase Form II

Both Form I and Form II of Formula (1) free base were processed undersimilar parameters using the process and composition for formulation F-2(as described above). Formula (1) was blended with the ingredients andthen lubricated, then roller compacted with a top feeding rollercompacter with a separate granulation step. The granules were thenlubricated. The resultant granules from the Form I and Form II were thencharacterized for tapped and aerated density. The Form II granulesshowed a general trend towards poor flow and poor uniformity.

Flowability typically affects the ease of handling pharmaceuticalproduct during processing. When flowability is very poor, problems occurwith handling and processing during blending, granulation andfilling/compression. The flowability based on interparticulateinteractions can be measured using the Hausner ratio or thecompressibility index by measuring the aerated and tapped density of thepowders. These values are calculated and ranked as outlined in the U.S.Pharmacoepia Monograph USP <1174> monograph. Hausner, Int. J. PowderMetall. 1967, 3, 7-13; Carr, Chem. Eng. 1965, 72, 163-168. The U.S.Pharmacoepia Monograph USP <1174> defines the following categories offlow character: Excellent (compressability index <10%, Hausner's ratio1.00 to 1.11); Good (compressability index 11-15%, Hausner's ratio 1.12to 1.18); Fair (compressability index 16-20%, Hausner's ratio 1.19 to1.25); Passable (compressability index 21-25%, Hausner's ratio 1.26 to1.34); Poor (compressability index 26-31%, Hausner's ratio 1.35 to1.45); Very poor (compressability index 32-37%, Hausner's ratio 1.46 to1.59); and Very, very poor (compressability index >38%, Hausner's ratio>1.60).

The Hausner ratio and compressibility index for the Form I granules was1.33 and 25% respectively while the Form II granules exhibited a Hausnerratio of 1.47 and compressibility index of 32%. The results thusindicates that Form I granules have a passable flow while Form IIgranules have poor to very poor flow.

The blends were then filled into capsules using an automatedencapsulator operating on the dosing disc principle. After filling to atarget weight, capsules are checked for weight uniformity with out ofweight capsules being rejected. The Form I capsules have a yield of90-100% acceptable capsules while the Form II containing capsules had ayield of only 40-60%.

Upon measuring the content uniformity as defined by the U.S.Pharmacoepia Monograph USP <905> for a hard gelatin capsule, the Form IIcontaining capsules have an acceptance value greater than 15, while theForm I containing capsules have an acceptance value below 15.

The results are summarized in Table 55.

TABLE 55 Results of Processability Tests for Form I and Form II ofFormula (1) free base. Bulk Density Tap Density Hausner CompressibilityLot g/cc g/cc ratio Index (%) F-2 using Form I 0.527 0.703 1.33 25 F-2using Form II 0.438 0.644 1.47 32

We claim:
 1. A composition comprising crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base.
 2. The composition of claim 1, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by a transmission X-ray powder diffractionpattern comprising peaks at 6.4, 8.6, 10.5, 11.6, and 15.7 °2θ±0.2 °2θ.3. The composition of claim 2, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by a transmission X-ray powder diffractionpattern further comprising peaks at 10.9, 12.7, 13.4, 14.3, 14.9, and18.2 °2θ±0.2 °2θ.
 4. The composition of claim 1, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by a transmission X-ray powder diffractionpattern further comprising one or more peaks selected from the groupconsisting of 11.3, 15.1, 15.7, 16.1, 17.3, 19.2, 19.4, 19.8, 20.7,21.1, 21.4, 21.6, 21.9, 22.6, 23.3, 23.6, 24.9, 25.2, 25.4, 25.7, 26.1,26.4, 26.8, 26.9, 27.7, 28.6, 29.1, 29.4, 30.1, 30.5, 31.7, 31.9, 32.2,32.6, 33.1, 33.4, 34.5, 35.9, 36.1, 36.8, 37.4, 38.1, 38.9, and 39.5°2θ±0.2 °2θ.
 5. The composition of claim 1, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by a transmission X-ray powder diffractionpattern substantially the same as the representative X-ray powderdiffraction pattern shown in FIG.
 2. 6. The composition of claim 1,wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by a Raman spectrum comprising peaks at 1620,1609, 1547, 1514 and 1495 cm⁻¹±4 cm⁻¹.
 7. The composition of claim 6,wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by a Raman spectrum further comprising one ormore peaks selected from the group consisting of 1680, 1620, 1609, 1574,1547, 1514, 1495, 1454, 1433, 1351, 1312, 1255, 1232, 1187, 1046, 995,706, 406, and 280 cm⁻¹±2 cm⁻¹.
 8. The composition of claim 1, whereinthe crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by a Raman spectrum substantially the same asthe representative Raman spectrum shown in FIG.
 2. 9. The composition ofclaim 1, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by an infrared (IR) spectrum comprising peaksat 1621, 1608, 1403, 1303, and 764 cm⁻¹±4 cm⁻¹.
 10. The composition ofclaim 9, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by an IR spectrum further comprising one ormore peaks selected from the group consisting of 3367, 3089, 2246, 1682,1621, 1608, 1574, 1514, 1504, 1454, 1428, 1403, 1345, 1303, 1248, 1194,1177, 1149, 1109, 1049, 1023, 1003, 947, 900, 858, 842, 816, 764, 734,729, 701, 689, 665, 623, and 612 cm⁻¹±4 cm⁻¹.
 11. The composition ofclaim 1, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is characterized by an IR spectrum substantially the same asthe representative IR spectrum shown in FIG.
 3. 12. The composition ofclaim 2, wherein the crystalline(S)-4-(8-amino-3-(1-(but-2-ynoyl)pyrrolidin-2-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamidefree base is further characterized by the absence of water in thecrystal structure.
 13. The composition of claim 1, further comprising anextragranular acidulant.
 14. The composition of claim 13, wherein theextragranular acidulant is selected from the group consisting of fumaricacid, succinic acid, D-tartaric acid, L-tartaric acid, racemic tartaricacid, ascorbic acid, isoascorbic acid (also known as erythorbic acid andD-araboascorbic acid), alginic acid, or a salt thereof, Protacid F 120NM, Protacid AR 1112 (also known as Kelacid NF), and Carbopol 971P(carboxypolymethylene), and combinations thereof.
 15. The composition ofclaim 13, wherein the extragranular acidulant is alginic acid, or asodium or potassium salt thereof, at a concentration of between about 5%to about 33% by weight.
 16. The composition of claim 1, wherein thecomposition further comprises at least one pharmaceutically-acceptableexcipient.
 17. A method of treating a hyperproliferative disease,comprising the step of administering a therapeutically effective amountof the composition of claim 21, wherein the hyperproliferative diseaseis selected from the group consisting of chronic lymphocytic leukemia,non-Hodgkin's lymphoma, diffuse large B-cell lymphoma, mantle celllymphoma, follicular lymphoma, B-cell lymphoproliferative disease, Bcell acute lymphoblastic leukemia, Waldenström's macroglobulinemia,Burkitt's leukemia, Hodgkin's disease, multiple myeloma, acute myeloidleukemia, juvenile myelomonocytic leukemia, hairy cell leukemia, mastcell leukemia, mastocytosis, myeloproliferative disorders (MPDs),myeloproliferative neoplasms, polycythemia vera (PV), essentialthrombocythemia (ET), primary myelofibrosis (PMF), myelodysplasticsyndrome, chronic myelogenous leukemia (BCR-ABL1-positive), chronicneutrophilic leukemia, chronic eosinophilic leukemia, primary centralnervous system (CNS) lymphoma, primary multifocal lymphoma of peripheralnervous system (PNS), thymus cancer, brain cancer, glioblastoma, lungcancer, squamous cell cancer, skin cancer (e.g., melanoma), eye cancer,retinoblastoma, intraocular melanoma, oral cavity and oropharyngealcancers, bladder cancer, gastric cancer, stomach cancer, pancreaticcancer, breast cancer, cervical cancer, head and neck cancer, renalcancer, kidney cancer, liver cancer, ovarian cancer, prostate cancer,colorectal cancer, bone cancer (e.g., metastatic bone cancer),esophageal cancer, testicular cancer, gynecological cancer, thyroidcancer, epidermoid cancer, AIDS-related cancer (e.g., lymphoma),viral-induced cervical carcinoma (human papillomavirus), nasopharyngealcarcinoma (Epstein-Barr virus), Kaposi's sarcoma, primary effusionlymphoma (Kaposi's sarcoma herpesvirus), hepatocellular carcinoma(hepatitis B and hepatitis C viruses), T-cell leukemias (Human T-cellleukemia virus-1), benign hyperplasia of the skin, restenosis, benignprostatic hypertrophy, tumor angiogenesis, chronic inflammatory disease,rheumatoid arthritis, atherosclerosis, inflammatory bowel disease, skindiseases such as psoriasis, eczema, and scleroderma, diabetes, diabeticretinopathy, retinopathy of prematurity, age-related maculardegeneration, hemangioma, ulcerative colitis, atopic dermatitis,pouchitis, spondylarthritis, uveitis, Behcet's disease, polymyalgiarheumatica, giant-cell arteritis, sarcoidosis, Kawasaki disease,juvenile idiopathic arthritis, hidratenitis suppurativa, Sjögren'ssyndrome, psoriatic arthritis, juvenile rheumatoid arthritis, ankylosingspondylitis, Crohn's disease, lupus, and lupus nephritis.
 18. The methodof claim 17, further comprising the step of administering atherapeutically effective amount of an acid-reducing agent.